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ArticleTransplantation Free access | 10.1172/JCI17477
Heterologous immunity provides a potent barrier to transplantation tolerance
Andrew B. Adams,1 Matthew A. Williams,1 Thomas R. Jones,1 Nozomu Shirasugi,1 Megan M. Durham,1 Susan M. Kaech,2 E. John Wherry,2 Thandi Onami,2 J. Gibson Lanier,2 Kenneth E. Kokko,1 Thomas C. Pearson,1 Rafi Ahmed,2 and Christian P. Larsen1
1Emory Transplant Center and Department of Surgery and 2Emory Vaccine Center and Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, Georgia, USA
Address correspondence to: Christian P. Larsen, Emory Transplant Center and Department of Surgery, Suite 5105 WMB, 1639 Pierce Drive, Atlanta, Georgia 30322, USA. Phone: (404) 727-8466; Fax: (404) 727-3660; E-mail: clarsen@emoryhealthcare.org.
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First published June 15, 2003 - More info
Published in Volume 111, Issue 12 on June 15, 2003
J Clin Invest. 2003;111(12):1887–1895. https://doi.org/10.1172/JCI17477.
First published June 15, 2003 - Version history
Received: November 26, 2002; Accepted: March 11, 2003
Many strategies have been proposed to induce tolerance to transplanted tissue in rodents; however, few if any have shown equal efficacy when tested in nonhuman primate transplant models. We hypothesized that a critical distinction between specific pathogen-free mice and nonhuman primates or human patients is their acquired immune history. Here, we show that a heterologous immune response — specifically, virally induced alloreactive memory — is a potent barrier to tolerance induction. A critical threshold of memory T cells is needed to promote rejection, and CD8+ "central" memory T cells are primarily responsible. Finally, treatment with deoxyspergualin, an inhibitor of NF-κB translocation, together with costimulation blockade, synergistically impairs memory T cell activation and promotes antigen-specific tolerance of memory. These data offer a potential explanation for the difficulty encountered when inducing tolerance in nonhuman primates and human patients and provide insight into the signaling pathways essential for memory T cell activation and function.
A critical feature of the adaptive immune response is the ability to recognize previously encountered pathogens. Memory T cells, in contrast to naive cells, are programmed to activate quickly, with a reduced requirement for costimulatory signals. Upon re-exposure to antigen, they rapidly regain effector function and generate a swift response that controls the pathogen and prevents disease (1). Phenotypic subsets of memory T cells, designated "central" and "effector" memory, have recently been described with distinct cytotoxic and proliferative capacities (2–4). However, the role of these individual subpopulations during secondary immune responses in vivo remains to be determined.
It is now clear that previous immunological exposures and resultant T cell memory can influence the course of future immune responses to unrelated pathogens, a phenomenon termed heterologous immunity (5). Considerably less is known regarding the effect of an individual's immune history on the response to an allogeneic tissue transplant. Although the frequency of naive T cells available to respond to any given pathogen is relatively small (approximately 1:200,000), the proportion that can directly recognize foreign MHC represents a substantial fraction of the total T cell repertoire (up to 1–10%) (6, 7). Given this extraordinary precursor frequency and the degeneracy of T cell recognition, it is not surprising that alloreactive T cells become activated after viral infection in experimental models (8–11). Similarly, allo-cross-reactivity for HLA-B14, -44, and -35 has been demonstrated by HLA-B8–restricted Epstein-Barr virus–specific (EBV-specific) T cells in humans (12). Interestingly, one of these cross-reactive combinations (B44 → B8) was identified as an "immunogenic pairing" (i.e., reduced allograft survival) in a renal transplant registry analysis (13). Further evidence of the clinical importance of cross-reactive memory T cells has been provided by Heeger et al., who have shown that a higher frequency of anti-donor memory T cells (presumably induced by prior environmental exposures) is associated with an increased rejection rate in clinical renal transplantation (14).
A variety of strategies have been devised to promote long-term allograft acceptance without the need for chronic immunosuppression in rodent models. Unfortunately, these protocols have proven to be markedly less effective when tested in preclinical primate models or in humans. One major distinction between specific pathogen-free mice and nonhuman primates or human patients is the exposure of the latter to pathogens and the resultant immune history. Although many tolerance strategies, such as regimens employing T cell costimulation blockade, successfully target and control naive T cell responses, memory cells have a lower threshold for activation (15, 16) and may not be susceptible. In support of this, Valujskikh et al. have shown that the presence of sensitized cells prevents the beneficial effects of anti-CD40L in prolonging allograft survival (17). Thus, strategies that successfully tolerize naive cells may fail when the host has pre-existing anti-donor memory, perhaps explaining the relative inability of these regimens to promote tolerance in large-animal models. Here, we provide evidence that an individual's immune memory can profoundly influence the outcome of tolerance induction protocols even when strategies are used that uniformly induce tolerance to highly immunogenic tissues in vigorous experimental models. First, we document that viral infections can lead to the generation of alloreactive memory T cells, which can confer resistance to tolerance induction. The quantity of accumulated anti-donor memory is critical — that is, there is a threshold of memory cells necessary to promote rejection — and we provide evidence that CD8+ central memory T cells are principally responsible for rejection. Further, there is a hierarchy in susceptibility to tolerance: naive animals are uniformly sensitive to tolerance induction; recipients exposed to a single prior infection show a slightly diminished susceptibility; and "immunologically experienced" hosts, having received multiple sequential infections, are refractory to tolerance induction. Finally, we show that the immunosuppressive agent 15-deoxyspergualin (DSG), an inhibitor of NF-κB translocation, synergizes with costimulation blockade to promote antigen-specific tolerance of both naive and memory T cells. These data suggest that heterologous immunity — in particular, virally induced alloreactive memory — may be an underappreciated barrier to tolerance induction in transplantation.
Mice and viral infections. BALB/c, C57BL/6 (B6, CD45.2), C57BL/6-Igh-6tm1Cgn, B6.SJL-Ptprca Pep3b/BoyJ (B6.SJL, CD45.1) mice were purchased from the Jackson Laboratory (Bar Harbor, Maine, USA). To induce acute infection, mice were inoculated with 2 × 105 PFUs of the lymphocytic choriomeningitis virus (LCMV) Armstrong strain (intraperitoneally), 5 × 106 PFUs of vaccinia virus (VV) (intraperitoneally), or 2 × 106 PFUs of vesicular stomatitis virus (VSV) (intravenously). For viral rechallenge experiments, immune mice received 2 × 106 PFUs of LCMV clone 13 (intravenously). Virus stocks were grown and quantitated as previously described (18). Infectious LCMV in serum and tissues was measured by plaque assay on Vero cell monolayers as described (18).
Skin grafting and tolerance induction protocols. Full-thickness skin grafts and the mixed chimerism tolerance protocol using CTLA4-Ig (Bristol-Myers Squibb, Princeton, New Jersey, USA), anti-CD40L (Bioexpress, Lebanon, New Hampshire, USA), and busulfan was administered as previously described (19). Rapamycin (0.4 mg/kg/day intraperitoneally; d0-6, Wyeth, Philadelphia, Pennsylvania, USA), anti–IL-2R mAb (250 μg intraperitoneally; d0,2,4,6-PC61), anti–common γ chain (500 μg intraperitoneally; d0,2,4,6-3E12/4G3, a gift from Thomas Malek), DSG (5 mg/kg per day intraperitoneally; and d0-6, a gift from Nippon Kayaku, Tokyo, Japan) were administered as indicated.
Cell preparations and flow cytometry. Intracellular IFN-γ expression was induced in response to 4–5 hours of ex vivo restimulation with LCMV peptides or allogeneic stimulators. In brief, splenocytes were prepared from experimental and naive animals. Responders were resuspended in cell culture media containing brefeldin (GolgiPlug, BD Pharmingen, San Diego, California, USA) and recombinant hIL-2 (10 ng/ml). To enumerate the allogeneic response, responders were plated in a 96-well plate at a 1:1 ratio (1 × 106 cells per well) with stimulators (splenocyte preparation from naive animals, either allogeneic [BALB/c] or syngeneic). To quantify the viral response, responder preparations were either incubated with viral peptides (LCMV and VSV) or with infected stimulator cells as previously described (20, 21). Responders alone, irrelevant peptide, or stimulation with syngeneic splenocytes were used as specificity controls. All stimulations were performed for 4–5 hours at 37°C. Intracellular staining for IFN-γ was performed as per the manufacturer's instructions (Cytofix/Cytoperm kit, BD Pharmingen). Peripheral blood was analyzed by staining with the indicated fluorochrome-conjugated antibodies (BD Pharmingen), followed by red blood cell lysis and washing with a whole-blood lysis kit (R&D Systems, Minneapolis, Minnesota, USA). Flow cytometry was performed on a FACSCalibur, and data were analyzed using Cellquest software (Becton-Dickinson, Braintree, Massachusetts, USA). Cell selections were achieved using a MACS magnetic sorting system (Miltenyi BioTec, Auburn, California, USA).
Statistical analyses. Statistical analyses were performed using a Mann-Whitney U test for skin graft survival; otherwise, a Student's t test was used.
Viral infections induce heterologous allospecific effector and memory responses. Although previous studies in both mice and man have documented allospecific cytotoxic T lymphocyte (CTL) activity immediately after viral infection (9–11), there is little quantitative information on the retention of alloreactive memory T cells after viral infections and their potential effects on transplantation. As an initial approach to determine whether prior exposure to viral pathogens might influence the response to transplanted allogeneic tissues, we studied the expansion and retention of virus-specific and alloreactive effector and memory cells during and after acute viral infections with LCMV (Armstrong strain), VV, or VSV. Each of these viruses produces an acute infection that stimulates an appropriate immune response, resulting in rapid clearance of the pathogen and the generation of a stable memory T cell population that persists for the life of the mouse (20, 22, 23). Mice (C57BL/6, H-2b) were infected with the individual viruses, and the number of virus-specific and alloreactive (H-2d–stimulated) IFN-γ–producing cells were measured in the effector phase (peak response) or during the memory phase (more than 6 weeks after infection) after viral clearance. For comparison, we measured the magnitude of the anti-donor T cell response during and after skin allografts (Figure 1). Age-matched naive control mice evaluated at similar time points never displayed detectable antiviral or anti–H-2d responses (data not shown).
Analysis of viral- and allo-specific responses in vivo. B6 mice were infected with either LCMV, VV, or VSV. The results are displayed for the peak of the response as well as during the memory phase (more than 6 weeks). For comparison, the response to a skin allograft is also shown. (a) Representative dot plots displaying the number of CD8+ IFN-γ+ T cells representing the antiviral response (LCMV-NP396-404, VV-infected stimulators, VSV-N-peptide; left panels) and the corresponding heterologous allo-specific responses (right panels). (b) The number of virus-specific (NP396-404, filled squares) and virally induced, alloreactive T cells (H-2d stimulated, filled triangles) after LCMV infection are shown over time. The allo-specific response after skin graft is shown for comparison (H-2d stimulated, open squares; CD8+, upper panel; CD4+, lower panel). Each time point represents the average for three animals. Experiments were repeated three times with similar results.
The number of virus-specific cells in both the effector and memory stages, as measured by intracellular staining for IFN-γ production (Figure 1a), was consistent with prior reports for each virus (20, 21, 24, 25). Animals infected with LCMV or VV generated a readily detectable heterologous anti–H-2d response, as measured by the number of T cells producing IFN-γ upon short-term, in vitro rechallenge with BALB/c splenocytes. As expected, the virally induced heterologous anti–H-2d response was lower in magnitude than the viral-specific response (LCMV [NP396-404], Figure 1b; VV [infected syngeneic cells], data not shown) or the anti-donor response after skin graft challenge (Figure 1b), but it still represented about 1–10% of the overall antiviral response. Despite previous reports that infection with VSV elicits a cross-reactive allogeneic response (anti–H-2bm8) (9), we found no comparable response to a distinct haplotype (H-2d), suggesting that the magnitude of the virally induced heterologous response is dependent on the particular virus-MHC pairing being evaluated.
Donor-specific memory prevents tolerance induction. Having demonstrated that an encounter with environmental pathogens results in the generation of memory T cells with specificity for foreign MHC, we next examined whether donor-reactive memory cells could prevent tolerance induction using a protocol that induces robust, life-long deletional tolerance to fully MHC-mismatched skin grafts (19). Briefly, mice received a short course of costimulation blockade (CTLA4-Ig and anti-CD40L) in addition to donor bone marrow infusion preceded by a single dose of busulfan, a stem cell–selective agent used to promote engraftment of the donor bone marrow cells. Naive mice treated with this protocol exhibit stable mixed hematopoietic chimerism (approximately 50% donor cells in peripheral blood) and deletional donor-specific tolerance (19).
B cell–deficient mice were initially chosen to study the effects of anti-donor memory on tolerance induction in order to permit the study of anti-donor T cell memory in the absence of anti-donor antibody. Although naive B6 B cell–deficient recipients (C57BL/6-Igh–/–) receiving the tolerance protocol developed stable mixed chimerism and accepted donor-specific BALB/c skin grafts (median survival time [MST], more than 75 days) (data not shown), sensitized (more than 60 days after primary BALB/c skin transplant) B cell–deficient recipients were refractory to tolerance induction and universally rejected their secondary donor allografts by three weeks (MST, 19 days) (data not shown). These results suggested that donor-specific memory T cells could abrogate the ability of the costimulation blockade–based protocol to induce mixed chimerism and tolerance.
A critical threshold of donor-specific memory T cells is required to resist tolerance induction. To ascertain whether a critical number of memory cells was needed to prevent tolerance induction, we developed an adoptive transfer system in which graded doses of T cells from sensitized wild-type B6.SJL (CD45.1) mice (skin grafted 60 days earlier) were transferred to naive congenic B6 (CD45.2) recipients before administration of the tolerance induction protocol. Before transfer, the number of donor-reactive CD4+ and CD8+ memory cells was quantified by intracellular staining for IFN-γ. Preliminary studies indicated that approximately 1% of CD4+ and approximately 2–4% of CD8+ splenic T cells possessed a memory phenotype (production of IFN-γ upon short-term in vitro restimulation [4–5 hours]). Two days after the transfer of cells, we then subjected the recipients to the tolerance induction protocol. Mice receiving T cells from naive congenic hosts were nearly uniformly (over 95%) susceptible to chimerism and tolerance induction. Resistance to tolerance was observed in recipients reconstituted with T cells purified from sensitized mice. Groups receiving lower doses of allo-specific memory cells followed a pattern similar to naive mice, with a slightly diminished susceptibility to tolerance induction. In contrast, the majority of mice receiving more than 5 × 106 whole T cells from sensitized hosts (more than 100,000 memory cells) failed to generate donor cell chimerism and rejected skin grafts by 1 month (Figure 2a).
A critical number of CD8+ central memory T cells are needed to resist tolerance. Naive B6 mice (CD45.2) received T cells from sensitized congenic B6 mice (CD45.1). (a) Titration of memory cell numbers. Naive recipients received varying doses of T cells (calculated number of memory cells transferred in brackets) from sensitized hosts (2 × 107 [400,000], filled squares; 107 [200,000], filled diamonds; 5 × 106 [100,000], filled triangles; 2 × 106 [40,000], open diamonds; 106 [20,000], open triangles; 2 × 107 naive cells [none], open squares). (b) CD8+ memory T cells represent a potent barrier to tolerance induction. Purified CD4+, CD8+, or whole T cells were isolated from sensitized mice and transferred to naive congenic mice. Mice receiving either naive selected cells (data not shown) or CD4+ T cells (filled diamonds) from sensitized donors developed high-level donor cell chimerism and accepted donor-type skin grafts indefinitely. The transfer of CD8+ T cells (filled triangles) from sensitized animals prevented tolerance induction in a similar fashion to transferred whole T cells (filled squares) (MST, 21 days). (c) CD8+ central memory T cells promote rejection more efficiently than "effector" memory T cells. Equal numbers of CD8+ (open squares), CD8+ CD62Lhi (filled squares), or CD8+ CD62Llo (open triangles) antigen-specific memory T cells were transferred to naive congenic hosts, which were then subjected to the tolerance protocol (control naive CD8+ T cells, filled triangles). Both unseparated CD8+ memory T cells and central CD8+ memory T cells efficiently rejected allogeneic skin grafts in less than 3 weeks (MST, 21 and 18 days, respectively). In contrast, effector memory T cells of the traditional CD62Llo memory phenotype were less effective at promoting rejection (MST, more than 70 days; P < 0.01).
CD8+ central memory T cells are the principal mediators of rejection. In additional studies, we investigated the relative contributions of CD8+ and CD4+ memory cells in promoting rejection. Equal numbers of purified CD4+ or CD8+ T cells from sensitized animals were transferred to naive congenic recipients before tolerance induction (CD4+ and CD8+ cell preparations were at least 90% pure with no more than 1% contamination of the complementary subset). Animals receiving CD8+ memory T cells quickly rejected their skin grafts, similar to animals receiving unfractionated memory cells (MST, 21 days). In stark contrast, animals that received CD4+ memory cells developed donor cell chimerism and enjoyed prolonged graft survival (more than 100 days) (Figure 2b). These data suggest that CD8+ memory cells represent a powerful barrier to tolerance induction in this model, and additional pathways, crucial for memory cell activation and function, need to be identified to allow for successful tolerance induction in the face of existing donor-specific CD8+ T cells.
Recently, there have been several reports documenting the phenotypic and functional heterogeneity that exists in the memory T cell compartment (2–4). A model of central and effector memory has been proposed because of differences in location and expression of homing molecules (CD62L and CCR7) (2). Central memory T cells (CD62Lhi CCR7+) lack immediate "killing" activity but rapidly produce IL-2 when restimulated in vitro, suggesting a greater proliferative capacity than their effector counterparts. Effector memory T cells (CD62Llo CCR7–), on the other hand, rapidly produce IFN-γ, contain intracellular perforin, and are capable of immediate ex vivo cytotoxicity. Given their capability for immediate effector function and favorable homing properties, it has been suggested that effector memory T cells would be better able to control subsequent immune challenges and that central memory T cells remain available in secondary lymphoid organs to provide "help" and to replenish dwindling effectors (1). In order to investigate the roles of effector and central memory T cells in preventing tolerance induction in our model, we transferred equal numbers (200,000) of antigen-specific (as assessed by IFN-γ production upon short-term restimulation in vitro) CD8+, CD8+ CD62Lhi, or CD8+ CD62lo memory T cells into naive congenic mice. We then subjected each group to the tolerance induction protocol as described above. As in previous experiments, mice receiving unfractionated CD8+ memory T cells quickly rejected donor allografts (MST, 21 days). Surprisingly, despite the lack of immediate cytotoxic potential, central CD8+ memory T cells promoted rejection more efficiently (MST, 18 days) than effector CD8+ memory T cells (MST, more than 70 days; P < 0.01) (Figure 2c).
Virally induced donor-specific memory cells prevent tolerance induction. Given the observations that viral infections induce alloreactive memory responses and that donor-specific memory can prevent tolerance, we established a model to evaluate the impact of virally induced allo-specific memory on tolerance induction. We hypothesized that although a single infection with LCMV may generate a sufficient heterologous response at the peak of the antiviral response to induce graft rejection, the number of alloreactive cells that remain in the memory phase may be insufficient to overcome the tolerance protocol. Furthermore, infection of a specific pathogen-free mouse with a single virus may not adequately simulate the clinical situation in which exposure to a myriad of pathogens and antigens is the rule and memory cells typically represent 40–50% of the T cell pool in adult humans (26). Thus, we studied animals that received multiple, distinct viral infections over time to more closely approximate the acquired immune history of human patients or outbred nonhuman primates, which commonly show evidence of exposure to many pathogens (e.g., EBV, CMV, VZV, polyoma).
Accordingly, we infected mice (C57BL/6) with either a single pathogen (LCMV, VV, or VSV) or sequentially with multiple pathogens (LCMV→VV or LCMV→VV→VSV), allowing 6–8 weeks between infections and at least 8 weeks after the final infection before study in the tolerance experiments to ensure that only memory cells remained. Uninfected, age-matched naive B6 mice uniformly developed high-level chimerism (mean chimerism, 49.1 ± 4.22 in 10 of 10 animals) and accepted donor skin grafts indefinitely (10 of 10; MST, more than 100 days) (Figure 3a). Mice that were immune to a single pathogen (LCMV, Figure 3, and VV, data not shown) demonstrated a slight decrease, although not statistically significant in this experiment, in the number of animals becoming chimeric and tolerant (9 of 12, P = 0.2 when compared with the naive group). This slightly diminished susceptibility to tolerance mirrored our earlier results when suboptimal doses of memory cells were transferred (Figure 2a). In other experiments, tolerance was induced in 37 of 45 LCMV-immune mice (82%), which is lower than the over 95% tolerance induction rate in naive mice, as observed in well over 200 nonconcurrent control recipients, suggesting that prior exposure to a potent infection such as LCMV may indeed have a moderate effect. In contrast to uninfected recipients, mice that had previously been exposed to multiple viral infections were refractory to tolerance induction and rejected their allografts (MST, 24 days; n = 12) (Figure 3a). In all groups, the presence of stable donor hematopoietic cell engraftment correlated with long-term skin graft survival (i.e., no chimerism indicated no skin graft survival) (Figure 3b). Not surprisingly, the number of virally induced alloreactive memory cells was higher in animals with multiple infections than in singly infected animals (data not shown).
Virally induced, alloreactive memory cells prevent tolerance induction. Naive, monoimmune (LCMV or VV), or polyimmune mice (LCMV→VV or LCMV→VV→VSV) received the mixed chimerism, costimulation blockade–based tolerance protocol. (a and b) Skin graft survival and donor chimerism levels after tolerance induction. Naive mice universally accept donor-type skin allografts (left panel, filled squares) and become high-level mixed chimeras (10 of 10, right panel). The majority of mice immune to a single pathogen become tolerant, although not as consistently as naive mice (9 of 12, open triangles). Mice immune to multiple pathogens are refractory to tolerance induction (3 of 12; MST, 24; P = 0.002 when compared with naive mice; filled diamonds). Inf, infection(s).
DSG, an inhibitor of NF-κB translocation, promotes tolerance of antigen-specific memory T cells. Although naive cells require stimulation through the T cell receptor and additional costimulatory signals for their activation, memory T cells are less stringent in their activation requirements. In an attempt to promote tolerance of antigen-specific memory T cells, we identified potential pathways that may inhibit memory cell activation. Anti-CD25, rapamycin, and blocking monoclonal antibodies against the common γ chain have all been reported to act synergistically with costimulation blockade to inhibit allograft rejection (27–29). In addition, we examined the effects of the novel immunosuppressant DSG, which has shown efficacy in both nonhuman primate models and human patients (30, 31). DSG is reported to bind to the intracellular chaperone Hsc70 and inhibit translocation of NF-κB, a transcription factor important for T cell activation and survival (32, 33).
Using our adoptive transfer model, we evaluated the agents for their ability to inhibit memory cell activation. Mice that had previously received T cells from sensitized congenic mice (approximately 200,000 memory T cells) were subjected to the tolerance protocol alone or the tolerance protocol in combination with the specific inhibitors. The addition of rapamycin, anti-CD25, or anti-γc failed to promote tolerance of anti-donor memory cells. The mice rejected their grafts at similar times as control animals receiving the tolerance protocol alone (Figure 4a), suggesting that memory cell activation and function was not critically dependent on those pathways. Interestingly, mice receiving the costimulation blockade–based tolerance protocol and DSG uniformly became tolerant and accepted their allografts indefinitely despite prior transfer of anti-donor memory cells (n = 5; MST, more than 100 days) (Figure 4a). Upon further follow-up, animals receiving the tolerance protocol and DSG accepted skin allografts and maintained stable donor chimerism for at least 1 year. When rechallenged with secondary donor and third-party skin allografts, animals accepted secondary donor skin grafts while rejecting third-party grafts in control time, suggesting that the combination of costimulation blockade and DSG effectively promoted tolerance of both naive and memory donor-reactive T cells (data not shown).
DSG and costimulation blockade synergistically inhibit memory cells. Both alloreactive and virus-specific memory cell responses were analyzed. (a) Skin graft survival of mice that had previously (1 week earlier) received T cells from sensitized congenic mice and were treated with either the tolerance induction protocol alone or in combination with various agents. The combination of DSG and costimulatory blockade promoted tolerance of donor-specific memory cells (n = 5; MST, 100 days; filled squares). The costimulation blockade tolerance regimen given alone (n = 5, open squares) or in combination with other agents, including rapamycin (n = 5, filled triangles), anti-CD25 (n = 5, open triangles), and anti-γc (n = 5, filled diamonds) failed to inhibit memory-dependent rejection. Two additional experiments demonstrated similar results. (b) LCMV-immune mice were rechallenged with LCMV clone 13 and treated with costimulation blockade (costim), DSG, or the combination of costimulation blockade and DSG. Five days after rechallenge, CD8+ (class I restricted NP396-404 and GP276-286) and CD4+ (class II restricted P13 GP60-80) responses were analyzed. The combination of costimulatory blockade and DSG synergistically inhibited LCMV-specific CD8+ memory T cells (*P < 0.01). No tx, no treatment; immune no rechall, immune mouse without viral rechallenge.
In order to more fully evaluate the effects of costimulation blockade and DSG on memory cell activation and function, we also studied recall responses in the LCMV model. Mice immune to LCMV (LCMV infection more than 60 days earlier) were rechallenged with LCMV and were administered costimulation blockade (CTLA4-Ig/anti-CD40L) alone, DSG alone, costimulation blockade and DSG, or no treatment. Five days after rechallenge, T cells were harvested and epitope-specific CD8+ (class I restricted NP396-404 and GP276-286) and CD4+ (class II restricted P13 GP61-80) memory responses were analyzed. Memory responses in mice treated with costimulation blockade were similar to those in immune control animals receiving no treatment (Figure 4b), consistent with studies demonstrating that memory T cells do not require costimulation delivered through CD28 or CD40L for their activation. DSG alone exerted only a moderate effect on the antiviral memory response, whereas the combination of costimulation blockade and DSG synergistically impaired the antiviral memory response — in particular, the CD8+ memory T cell response (Figure 4b) (P = 0.01 when compared with no treatment).
To determine if the combination of costimulation blockade and DSG promoted tolerance of memory cells in an antigen-specific manner, we transferred memory cells from LCMV-immune mice (approximately 500,000 P14 memory T cells) and from allo-immune mice (approximately 400,000 donor-specific memory T cells). Two days later, we administered the tolerance protocol alone or in combination with DSG (Figure 5a). Similar to previous results, mice receiving the tolerance protocol and DSG accepted donor-specific skin grafts, indicating effective inhibition of the transferred allo-memory cells. Animals receiving the tolerance protocol alone rejected grafts by 1 month (n = 5; MST, 28 days) (data not shown). To test whether the effect of DSG was antigen specific, we assessed the functional response of the transferred memory cells in both groups 2 months after transplantation. As expected, mice treated with the protocol and DSG failed to display any donor-reactive T cells (Figure 5b), indicating that the regimen had effectively tolerized the transferred allo-specific memory cells. Conversely, mice treated with the costimulation blockade tolerance protocol alone clearly exhibited the presence of donor-reactive memory cells (Figure 5b). All of the IFN-γ+ cells resided in the transferred population (CD45.1+) (data not shown), suggesting that the tolerance protocol effectively inhibited the generation of new effectors but failed to control pre-existing memory cells. Importantly, both groups displayed similar numbers of functional, virus-specific memory T cells, indicating that the combination of costimulation blockade and DSG acts synergistically to impair memory T cell responses in an antigen-specific manner, whereas pre-existing memory cells, which are not engaged by their cognate antigen during the treatment period, are preserved for future challenges.
Antigen-specific tolerance of memory cells. (a) Congenic mice were either infected with LCMV (Thy 1.1) or received a skin allograft (CD45.1). Eight weeks after challenge, T cells were isolated to obtain both allo- and virus-specific memory cells and then transferred into naive B6 mice (CD45.2/Thy1.2). Mice then received the tolerance protocol or the tolerance protocol with DSG. sg, skin graft. (b) When the tolerance protocol alone was administered, both alloreactive and virus-specific memory cell populations were preserved, resulting in allograft rejection. When given in combination with DSG, donor-reactive cells were specifically deleted, and viral memory remained intact upon rechallenge. Unstim, unstimulated. (c) Model for heterologous immunity as a barrier to transplantation tolerance.
Memory is an indispensable component of the immune response. The primary purpose of the amnestic response is to protect the host from future challenges of the identical pathogen. There is also accumulating evidence to suggest that in certain settings, memory elicited by certain viruses can enhance the clearance of even unrelated pathogens (5). Unfortunately, the heterologous response may also lead to immune-mediated disease in some circumstances (34, 35). Another potentially adverse consequence of an antiviral response is the production of effector T cells capable of directly recognizing foreign MHC and the subsequent formation of long-lived alloreactive memory cells.
An unresolved area of intense interest in the field of transplantation biology has been the somewhat puzzling and recurring observation that tolerance strategies, which promote long-lived graft survival in rodents, have consistently failed when evaluated in nonhuman primate models and clinical trials. This has led to the emerging concept that environmental exposures and large memory pools may constitute a more formidable barrier to tolerance induction than the immune system of experimental rodents.
Although cross-reactivity of pathogen-reactive T cells with allogeneic MHC complexes has been demonstrated at the cellular level, our results provide direct in vivo evidence that prior immune encounters can provide a powerful barrier to tolerance induction. Viral infections lead to the generation of alloreactive memory T cells, which in turn confer resistance to tolerance induction. Whether this is the result of bystander activation, mimicry on a molecular level (viral peptide bound by self-MHC resembling self-peptide bound by foreign MHC) or a combination of the two remains to be fully elucidated (36, 37). The number of memory cells the host has accumulated is critical; as we have shown, there is a threshold of memory cells necessary to promote rejection. Although naive animals and those receiving a single infection were susceptible to tolerance induction, it is very likely that these animals have encountered far fewer pathogens than potential transplant recipients, in whom the prevalence of viral infections (including EBV, CMV, HSV, VSV, influenza, etc.), as in the general population, is quite high. In support of our hypothesis, we have shown that animals immune to multiple viruses, perhaps a closer approximation to "immune-experienced" hosts, almost universally fail to become tolerant when using a regimen that uniformly leads to tolerance in naive recipients.
Our results indicate that CD8+ memory T cells are the principal mediators of rejection in this model. However, this does not necessarily exclude a role for CD4+ memory as a barrier to tolerance induction in other settings. In this model, the number of CD4+ memory T cells generated in vivo and then transferred to naive hosts was approximately fourfold lower than the number of memory CD8+ T cells. Thus, in this system, the number of CD4+ memory T cells may not achieve the critical threshold needed to precipitate rejection.
Perhaps one of the most interesting new areas of investigation in immunology is the description of functional subsets within the memory T cell pool. Although the properties of these subsets have not yet been completely defined, one might have anticipated that effector memory cells, which are programmed to home to peripheral locations and primed for immediate effector function, would constitute a more formidable barrier to tolerance than their central memory counterparts. Interestingly, our data indicate that central memory T cells are actually considerably more potent on a per-cell basis in their ability to resist tolerance induction. This suggests that central and effector memory may differ in dependence of each on CD28 and CD40 costimulatory signals or that the superior proliferative capacity of central memory cells is necessary to resist tolerance.
Although the signal requirements for memory cell activation and function have not been completely defined, traditional costimulatory signals (e.g., CD28/B7, CD40/CD40L) do not seem to be essential (15, 16). Recent work has identified cytokines (IL-15 and IL-7) important for maintenance of memory through homeostatic proliferative mechanisms (35, 38–40). Interestingly, neither of these cytokines was essential for productive recall responses. Consistent with these data, we present data to suggest that the interruption of key cytokine signaling pathways, using either blocking antibodies (anti-IL-2R or anti-γc) or the immunosuppressive molecule rapamycin, did not significantly inhibit memory cell function in our model. In contrast, the combination of T cell costimulation blockade and the immunosuppressive agent DSG synergized to inhibit the function of memory T cells in both allograft and viral models. The putative mechanism of action of DSG is to inhibit translocation of NF-κB, a transcription factor involved in many cellular processes, including T cell activation (41). Interestingly, NF-κB plays a critical role in determining the balance between pro- and antiapoptotic factors, which in turn determine the fate of the cell (42, 43). The experiments in which we cotransfer LCMV-specific and allo-specific memory cells indicate that the combination of DSG and costimulation blockade can induce inactivation of antigen-specific memory cells, since only those cells encountering their cognate ligands during the period of drug administration were affected. Protective immunity was preserved, since viral memory cells not specific for the tolerogen (donor marrow) remained fully competent and capable of responding to a subsequent viral rechallenge. Inhibition of NF-κB at a key step in memory T cell activation may result in a decrease in essential cellular survival signals and tip the balance toward cell death, permitting the deletion of antigen-specific memory cells.
Our data suggest that heterologous immunity, in particular virally induced alloreactive memory, may represent an underexplored barrier to tolerance induction in transplantation. It provides a potential explanation for the lack of success when tolerance regimens, effective in rodent models, are tested in nonhuman primate models or in human patients. Although cross-reactive responses between unrelated viruses have been documented, simple probability suggests that the occurrence of heterologous allogeneic responses after pathogen exposure should be even more likely (given the high precursor frequency of the direct allogeneic response).
There are three distinct possibilities whereby pathogen exposure could influence the host's response to allo-antigen. First there may be an active, ongoing infection at the time of tolerance induction. This includes exposure to an acute infection coincidental with attempts at tolerance or the presence of a smoldering persistent, productive infection in which the pathogen is present but under immune control. Potential unfavorable outcomes include the possibility of tolerance to the pathogen and loss of immune control, or the ongoing antiviral response may promote rejection and an inability to induce tolerance. We and others have attempted to address this question experimentally and in prior studies have demonstrated that a concurrent acute infection with LCMV at the time of tolerance induction can precipitate rejection (36, 37). Recently, we have also demonstrated that the presence of an ongoing, persistent productive viral infection interferes with attempts to induce tolerance (44). Heeger and colleagues have recently published similar findings for the chronic parasitic infection Leishmania (45). Both of these studies address a similar question in systems in which there is eventual immune control of the infection but the pathogen is never completely cleared and persists for the life of the mouse (18, 46, 47). This may be of particular relevance to a subset of transplant patients who suffer from ongoing, chronic infections such as hepatitis C. However, mechanisms in addition to simple cross-reactivity may also be important in these models. For example, the active immune response against the persistent pathogen may in and of itself provide a means, such as elevated levels of cytokines and growth factors, to overcome costimulatory blockade.
The second scenario is exposure of the host to multiple acute viral infections that resolve completely before attempts of tolerance are initiated. This condition, which is present in virtually all human transplant recipients, has not previously been investigated experimentally. In this work, we examined the influence of prior resolved infections on the host's memory pool and demonstrated dramatic effects on tolerance induction. Our results indicate that a single prior acute viral infection (e.g., LCMV, VV), which is subsequently completely cleared, is inadequate to generate a sufficient number of alloreactive memory T cells necessary to reach the critical threshold required to impair attempts at tolerance and promote rejection. Humans, unlike experimental rodents, have been exposed to a wide variety of pathogens, each of which has the potential to generate alloreactive memory (Figure 5c). Our data indicate that if sufficient numbers of donor-specific memory T cells accumulate after multiple acute viral infections over time, a critical threshold is exceeded, and attempts at tolerance induction that fail to target memory cells will be unsuccessful (Figure 5c). Lastly, the influence of pathogens that are harbored in a quiescent, true latent state has yet to be tested experimentally. Ongoing studies in our laboratory seek to address this important issue.
In conclusion, while further work is necessary to identify the pathways critical for memory cell activation and survival, the combination of costimulation blockade and DSG offers a clinically relevant strategy to promote antigen-specific tolerance of environmentally induced, alloreactive memory T cells. Thus, future plans for tolerance trials in humans should take into consideration that even recipients of primary transplants may not be as "naive" as practitioners believe.
This work was supported in part by research grants DK/AI40519, CA74364-03, and AI44644 from the National Institutes of Health; by the Engineering Research Center Program of the National Science Foundation under award number EEC-9731643; and by the Carlos and Marguerite Mason Trust.
See the related Commentary beginning on page 1819.
Nonstandard abbreviations used: Epstein-Barr virus (EBV); 15-deoxyspergualin (DSG); lymphocytic choriomeningitis virus (LCMV); vaccinia virus (VV); vesicular stomatitis virus (VSV); cytotoxic T lymphocyte (CTL); median survival time (MST).
Kaech, SM, Wherry, EJ, Ahmed, R. Effector and memory T-cell differentiation: implications for vaccine development. Nat. Rev. Immunol. 2002. 2:251-262.
Sallusto, F, Lenig, D, Forster, R, Lipp, M, Lanzavecchia, A. Two subsets of memory T lymphocytes with distinct homing potentials and effector functions. Nature. 1999. 401:708-712.
Masopust, D, Vezys, V, Marzo, AL, Lefrancois, L. Preferential localization of effector memory cells in nonlymphoid tissue. Science. 2001. 291:2413-2417.
Reinhardt, RL, Khoruts, A, Merica, R, Zell, T, Jenkins, MK. Visualizing the generation of memory CD4 T cells in the whole body. Nature. 2001. 410:101-105.
Welsh, RM, Selin, LK. No one is naive: the significance of heterologous T-cell immunity. Nat. Rev. Immunol. 2002. 2:417-426.
Blattman, JN, et al. Estimating the precursor frequency of naive antigen-specific CD8 T cells. J. Exp. Med. 2002. 195:657-664.
Suchin, EJ, et al. Quantifying the frequency of alloreactive T cells in vivo: new answers to an old question. J. Immunol. 2001. 166:973-981.
Mason, D. A very high level of crossreactivity is an essential feature of the T-cell receptor. Immunol. Today. 1998. 19:395-404.
Sheil, JM, Bevan, MJ, Lefrancois, L. Characterization of dual-reactive H-2Kb-restricted anti-vesicular stomatitis virus and alloreactive cytotoxic T cells. J. Immunol. 1987. 138:3654-3660.
Yang, H, Welsh, RM. Induction of alloreactive cytotoxic T cells by acute virus infection of mice. J. Immunol. 1986. 136:1186-1193.
Braciale, TJ, Andrew, ME, Braciale, VL. Simultaneous expression of H-2-restricted and alloreactive recognition by a cloned line of influenza virus-specific cytotoxic T lymphocytes. J. Exp. Med. 1981. 153:1371-1376.
Burrows, SR, Khanna, R, Burrows, JM, Moss, DJ. An alloresponse in humans is dominated by cytotoxic T lymphocytes (CTL) cross-reactive with a single Epstein-Barr virus CTL epitope: implications for graft-versus-host disease. J. Exp. Med. 1994. 179:1155-1161.
Maruya, E, Takemoto, S, Terasaki, PI. HLA matching: identification of permissible HLA mismatches. Clin. Transpl. 1993. 9:511-520.
Heeger, PS, et al. Pretransplant frequency of donor-specific, IFN-gamma-producing lymphocytes is a manifestation of immunologic memory and correlates with the risk of posttransplant rejection episodes. J. Immunol. 1999. 163:2267-2275.
Lanzavecchia, A, Sallusto, F. Dynamics of T lymphocyte responses: intermediates, effectors, and memory cells. Science. 2000. 290:92-97.
London, CA, Lodge, MP, Abbas, AK. Functional responses and costimulator dependence of memory CD4+ T cells. J. Immunol. 2000. 164:265-272.
Valujskikh, A, Pantenburg, B, Heeger, PS. Primed allospecific T cells prevent the effects of costimulatory blockade on prolonged cardiac allograft survival in mice. Am. J. Transplant. 2002. 2:501-509.
Ahmed, R, Salmi, A, Butler, LD, Chiller, JM, Oldstone, MB. Selection of genetic variants of lymphocytic choriomeningitis virus in spleens of persistently infected mice. Role in suppression of cytotoxic T lymphocyte response and viral persistence. J. Exp. Med. 1984. 160:521-540.
Adams, AB, et al. Costimulation blockade, busulfan, and bone marrow promote titratable macrochimerism, induce transplantation tolerance, and correct genetic hemoglobinopathies with minimal myelosuppression. J. Immunol. 2001. 167:1103-1111.
Harrington, LE, Whitton, JL, Ahmed, R. Recombinant vaccinia virus-induced T-cell immunity: quantitation of the response to the virus vector and the foreign epitope. J. Virol. 2002. 76:3329-3337.
Murali-Krishna, K, et al. Counting antigen-specific CD8 T cells: a reevaluation of bystander activation during viral infection. Immunity. 1998. 8:177-187.
Lau, LL, Jamieson, BD, Somasundaram, T, Ahmed, R. Cytotoxic T-cell memory without antigen. Nature. 1994. 369:648-652.
Kundig, TM, et al. On the role of antigen in maintaining cytotoxic T-cell memory. Proc. Natl. Acad. Sci. U. S. A. 1996. 93:9716-9723.
Whitmire, JK, Ahmed, R. Costimulation in antiviral immunity: differential requirements for CD4(+) and CD8(+) T cell responses. Curr. Opin. Immunol. 2000. 12:448-455.
Butz, EA, Bevan, MJ. Massive expansion of antigen-specific CD8+ T cells during an acute virus infection. Immunity. 1998. 8:167-175.
Douek, DC, et al. Changes in thymic function with age and during the treatment of HIV infection. Nature. 1998. 396:690-695.
Li, Y, et al. Blocking both signal 1 and signal 2 of T-cell activation prevents apoptosis of alloreactive T cells and induction of peripheral allograft tolerance. Nat. Med. 1999. 5:1298-1302.
Jones, TR, et al. The role of the IL-2 pathway in costimulation blockade-resistant rejection of allografts. J. Immunol. 2002. 168:1123-1130.
Demirci, G, et al. On CD28/CD40 ligand costimulation, common gamma-chain signals, and the alloimmune response. J. Immunol. 2002. 168:4382-4390.
Bartholomew, AM, et al. Tolerance in a concordant nonhuman primate model. Transplantation. 1999. 68:1708-1716.
Knechtle, SJ. Treatment with immunotoxin. Philos. Trans. R. Soc. Lond. B Biol. Sci. 2001. 356:681-689.
Nadler, SG, Tepper, MA, Schacter, B, Mazzucco, CE. Interaction of the immunosuppressant deoxyspergualin with a member of the Hsp70 family of heat shock proteins. Science. 1992. 258:484-486.
Tepper, MA, Nadler, SG, Esselstyn, JM, Sterbenz, KG. Deoxyspergualin inhibits kappa light chain expression in 70Z/3 pre-B cells by blocking lipopolysaccharide-induced NF-kappa B activation. J. Immunol. 1995. 155:2427-2436.
Selin, LK, Varga, SM, Wong, IC, Welsh, RM. Protective heterologous antiviral immunity and enhanced immunopathogenesis mediated by memory T cell populations. J. Exp. Med. 1998. 188:1705-1715.
Chen, HD, et al. Memory CD8+ T cells in heterologous antiviral immunity and immunopathology in the lung. Nat. Immunol. 2001. 2:1067-1076.
Tan, JT, et al. Interleukin (IL)-15 and IL-7 jointly regulate homeostatic proliferation of memory phenotype CD8+ cells but are not required for memory phenotype CD4+ cells. J. Exp. Med. 2002. 195:1523-1532.
Welsh, RM, et al. Virus-induced abrogation of transplantation tolerance induced by donor-specific transfusion and anti-CD154 antibody. J. Virol. 2000. 74:2210-2218.
Williams, MA, et al. Characterization of virus-mediated inhibition of mixed chimerism and allospecific tolerance. J. Immunol. 2001. 167:4987-4995.
Goldrath, AW, et al. Cytokine requirements for acute and basal homeostatic proliferation of naive and memory CD8+ T cells. J. Exp. Med. 2002. 195:1515-1522.
Becker, TC, et al. Interleukin 15 is required for proliferative renewal of virus-specific memory CD8 T cells. J. Exp. Med. 2002. 195:1541-1548.
Schluns, KS, Williams, K, Ma, A, Zheng, XX, Lefrancois, L. Cutting edge: requirement for IL-15 in the generation of primary and memory antigen-specific CD8 T cells. J. Immunol. 2002. 168:4827-4831.
Karin, M, Lin, A. NF-kappaB at the crossroads of life and death. Nat. Immunol. 2002. 3:221-227.
Beg, AA, Baltimore, D. An essential role for NF-kappaB in preventing TNF-alpha-induced cell death. Science. 1996. 274:782-784.
Williams, MA, et al. Cutting edge: Persistent viral infection prevents tolerance induction and escapes immune control following CD28/CD40 blockade-based regimen. J. Immunol. 2002. 169:5387-5391.
Pantenburg, B, Heinzel, F, Das, L, Heeger, PS, Valujskikh, A. T cells primed by Leishmania major infection cross-react with alloantigens and alter the course of allograft rejection. J. Immunol. 2002. 169:3686-3693.
Reiner, SL, Locksley, RM. The regulation of immunity to Leishmania major. Annu. Rev. Immunol. 1995. 13:151-177.
Sacks, D, Noben-Trauth, N. The immunology of susceptibility and resistance to leishmania major in mice. Nat. Rev. Immunol. 2002. 2:845-858.
Version 1 (June 15, 2003): No description

redpajama_set_name: RedPajamaCommonCrawl


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text:
Q: Allow user to click a button only when email is valid I am using angularjs on a web application. I am trying to perform something quite simple. I have an email input and a button. I only want the button's function to be enabled once a valid email is entered. I've tried:

*

    <input type="email" ng-model="email" placeholder="Enter Email" required>

Go!

Be default, i'd like the button to be blue. But once a valid email is entered, then the button turns Green. Hence the:
ng-class={true : 'validEmail', false : 'invalidEmail'}[email]

A: Here you are:
 ng-class="myForm.input.$valid ? 'valid' : 'invalid'"

ng-class checks whether the input is valid or invalid and returns two corresponding classes for .valid and .invalid.
While then you can apply styles with CSS:
.valid {
  background: green;
}

.invalid {
  background: blue;
}

The HTML should be something like this:
<form name="myForm" ng-controller="ExampleController">
  Email:
    <input type="email" name="input" ng-model="email.text" required>

        Required!

        Not valid email!

  <tt>text = {{email.text}}</tt>

  <button ng-class="myForm.input.$valid ? 'valid' : 'invalid'">Submit</button>
</form>

In controller:
var app = angular.module('plunker', []);

app.controller('ExampleController', ['$scope', function($scope) {
  $scope.email = {
    text: 'me@example.com'
  };
}]);

Also you can use ng-disabled="myForm.input.$invalid" on the button to make it unusable if the input is invalid.
See an Example.
Resources: input[email], The many ways to use ngClass, ngDisabled.

redpajama_set_name: RedPajamaStackExchange


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It is a rare pleasure when an entrepreneur approaches an Intellectual Property (IP) Attorney with concurrent requests for trademark (TM) and prior art searches early on. Both types of searches can save avoidable costs. But, this is not to suggest that you slam the brakes on the train before you get their results.
First, the name of your company might change during your marketing efforts. In the meantime, you can usually use a working name without seriously impeding your efforts. But, keep records of what names and any advertising, packaging, etc. that you use. Hold off committing serious money to such advertising, though, until you receive the results of the search.
Patenting operates on a different time table. The time between where you are and your (potential) patent has two primary road hazards. One, the public disclosure bar to patentability, can be avoided by keeping your discussions under non disclosure agreements (NDA); by marking all of your documents, prototypes, etc. with proprietary warnings; and by obtaining verbal confirmation of confidentiality before discussing your idea with others.
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A sneak peek at how organisers are curating next year's events.
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This rotting wooden goblet is filled with a festering brew of pus and blood, and writhing maggots and worms spill from the froth on the liquid's surface. A Shaman who sups from the chalice feels a burning pain wrack his body as the foul liquid seeps through him, yet his magical ability is suddenly enhanced, making his spells all-powerful and deadly.
The Shaman may drink from the Chalice at the start of his Magic phase. He suffers an immediate Strength 4 hit that ignores Armour and Ward saves (Regeneration may be taken as normal). For the remainder of that Magic phase, any spell cast by the Shaman will be cast with irresistible Force on a successful casting roll which includes and double, except a Miscast.


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text:
using System;
using System.Runtime.InteropServices;

namespace Esthar.Core
{
    [AttributeUsage(AttributeTargets.Class | AttributeTargets.Struct)]
    public sealed class UnsafeCastContainsOnlyValueTypesAttribute : Attribute
    {
    }

    public static class ArrayExm
    {
        public static T[] UnsafeInplaceCast<T>(this byte[] array)
        {
            unsafe
            {
                fixed (void* ptrArray = array)
                {
                    ArrayHeader* ptrHeader = (ArrayHeader*)ptrArray - 1;

                    ptrHeader->Length = (IntPtr)(array.Length / TypeCache<T>.UnsafeSize);
                    ptrHeader->TypeHandle = TypeCache<T[]>.Type.TypeHandle.Value;
                }
            }

            return (T[])(object)array;
        }

        [StructLayout(LayoutKind.Sequential, Pack = 1)]
        private struct ArrayHeader
        {
            public IntPtr TypeHandle;
            public IntPtr Length;
        }
    }
}

redpajama_set_name: RedPajamaGithub


text:
Николай Крумов Боев (8 май 1922 – 12 ноември 1985) е български зоолог, орнитолог и природозащитник

Произход
Николай Крумов Боев е роден на 8 май 1922 г. в Айтос, Бургаска област. Баща му, Крум Боев е книжар и издава вестника "Нов Айтос". Майка му, Златка Георгиева Боева е учителка по френски и руски език в прогимназията и гимназията в града и основателка на Българо-съветското дружество във Варна. Единият му дядо – Никола Боев е опълченец, участвал в битката при Стара Загора, а другият – войводата Дико Джелебов е български революционер, войвода на Вътрешната македоно-одринска революционна организация, първият знаменосец на Илинденско-Преображенското въстание. В семейството са три деца – има брат Милко и сестра Димка.

Образование
Основното и прогимназиалното си образование завършва в Айтос. В отделенията е ученик в класа на учителя Никола Стаматов, известен със своята взискателност. От малък Николай проявява голям интерес към животните. По това време прави своите първи системни наблюдения върху птиците в Айтос и събира лична сбирка от насекоми, която наброява няколкостотин вида.

Гимназиалното си образование завършва в Първа мъжка гимназия през 1941 г. във Варна. През този период интересът му към изучаването на животните се засилва. Както сам пише за себе си, със зоология се занимава още от ученическата скамейка. В гимназията и по-късно в Софийския университет главното му влечение остават зоологията и по-специално – птиците. Като ученик отглежда вкъщи някои по-обикновени видове птици, земноводни, влечуги и бозайници. Едва 9-годишен той си доставя и жадно прочита книгата на Едуард Клайн – най-пълното за времето си съчинение по птиците в България, съдържащо данни за всичките 314 известни дотогава в страната видове птици. Зоологът Иван Василев му е учител по биология в VII клас, а в VIII клас и класен наставник и това може да е оказало някакво положително въздействие върху дооформянето на интересите му.

Животинският свят и зоологията, обикнати още в детските години, задържат интереса му и като гимназист, когато вече познава почти всички български видове птици. По това време той редовно следи каталозите на различни книжарници и издателства и се снабдява с редица ценни зоологични издания за птици и бозайници. Първата си статия "Прокълнати птици"  написва още като ученик на 18-годишна възраст през 1941 г. Изпраща я в авторитетното списание "Природа" на Константин Христович (син на основателя му, Георги Христович и докато е в армията, статията му бива отпечатана (1942). Там излиза и втората му статия (1944). . Баща му иска Николай да стане търговец, защото той и баща му му също са били търговци (книжари) и е против продължаването на обучението му в гимназията. След настояване от страна на майка му, Николай заминава за Варна. Още като ученик (в VII клас) започва да събира народни имена на животните за съставянето на един бъдещ "Български зоологичен речник" – една необходимост и негова юношеска мечта, останала неосъществена до края на живота му, макар че издава един въпросник по темата

Като войник топограф-фотограметър през 1941 – 1943 г. в Шумен) Николай Боев събира вкаменелости из Шуменското плато и чете редица произведения на италиански, френски, немски, руски и английски учени-естественици, както и някои съчинения на проф. Асен Златаров. Докато е в армията, за кратко време е командирован от Шумен в София. Отпуските си прекарва в Царския Естествено-исторически музей, където почти всяка събота и неделя опитва с наличните си книги да определя птици и бозайници, подредени във витрините. Там се запознава лично и с Цар Борис III – познавач на птиците, които, подобно на баща му, били едно от неговите хобита. Още като войник, Николай Боев е в интензивна кореспонденция с един от водещите по онова време орнитолози в света – италианеца Едгардо Молтони Edgardo Moltoni, който му изпраща всички свои научни публикации. След 1944 г. контактите на Н. Боев с Молтони били са невъзможни, а мечтите да продължи образованието си в Италия – разбити завинаги.

Ранна научна дейност
През 1943 г. започва следването си по естествена история в Природоматематическия факултет на Софийския университет "Св. Климент Охридски" (1943 – 1947 г.). За началото на следването си той пише: ... Но почнаха бомбардировките и едва изкарахме първия семестър. За целия втори семестър бяхме разпуснати. През това време писах едно обширно писмо до Павел Патев (тогава директор на Зоологическата градина и единствения наш орнитолог), с което му пратих списък на нови за нашата фауна 30 вида птици според литературни данни и проучванията ми от сбирката птици в музея (като войник). През целия този свободен от занятия втори семестър аз правех наблюдения върху птиците на гр. Айтос, четях специална литература ..." В обширното си писмо-отговор, П. Патев потвърждава неговите издирвания по отношение на състава на нашите птици и му благодари за някои нови данни, които бил пропуснал. Патев му изпратил и пръстени за маркиране на птиците, с които Николай опръстенил стотина лястовици. По-късно Патев включва в монографията си "Птиците в България" (1950) данните от неговия "Списък на нови за нашата страна 30 вида птици", което е и първият сериозен научен принос на Боев в изучаването на българската орнитофауна.

С Павел Патев обаче Николай се запознава лично едва през 1945 г. като студент второкурсик. Тогава двамата със своя приятел и колега, бележитият наш зоолог, ст.н.с. Димо Божков, отиват в кабинета му и взаимно очаровани от посещението си, поставят началото на цяла серия срещи с Патев, от когото се научили да препарират и определят птиците и се запознали с много важни въпроси на орнитологията. Както отбелязва Д. Божков, Патев "... бързо оцени дарбите и способностите на Николай. Неслучайно по-късно той го привлече да му помага при подреждането на орнитологичните сбирки ..."
През студентските си години Н. Боев се заема с опръстеняването на птици с увлечение. Той е доброволен сътрудник на Българска орнитологическа централа при Царския Естествено-историческия музей в София. Като студент във II курс д-р Иван Буреш му предлага да подреди орнитологичните сбирки в музея след бомбардировките през март 1944 г. Тази задача за него е въпрос на чест и той я изпълнява блестящо "... безплатно и с удоволствие." И по-късно като лаборант, Н. Боев доброволно помага за подреждането на колекциите от гръбначни животни (и особено от птици). Първоначално това става под ръководството на акад. д-р Иван Буреш и Павел Патев, когото нарича "моя учител". Силно пострадалите от бомбардировките сбирки се нуждаели от реставриране, почистване и подреждане, което Боев прави с желание и интерес. Тази дейност съвпада с цялостното ремонтиране на музея, организирано и проведено от д-р Кръстю Тулешков и по-късно довършено от д-р Нено Атанасов. По политически съображения след 9 септември 1944 г. за Николай Боев настъпват трудни години. Той е лишен от право на стипендия и е принуден да следва и завърши образованието си на собствена издръжка.

Като студент, по повечето дисциплини Николай показва задълбочени знания и отличен успех. Почти всички от преподавателите му, видни наши естественици (проф. Георги Паспалев, проф. Николай Стоянов, проф. Даки Йорданов, доц. Пенчо Дренски, доц. Александър Вълканов, проф. Петър Бакалов), са възхитени от знанията на Николай. През тези години по молба на свои колеги, Николай често пише лозунги и рисува плакати, стенвестници и агиттабла и така участва активно в различни студентки прояви. Д-р Д. Костов, си спомня: "Кольо още от малък имаше тази природна дарба да рисува много хубаво. За възрастта му това бяха отлични рисунки." Тази му способност е оценена и от Павел Патев, който му предлага да изготви щриховите рисунки в монографията му за българските птици – задача, която Н. Боев блестящо изпълнява. През този период при всяка възможност младият Николай трупа знания, почерпени от неговите преподаватели и по-възрастни колеги в столицата. Заедно с това вече публикува и редица статии по интересни за него теми. Като студент отпечатва 20 научно-популярни статии, а до 25-ата си година има вече над 40 статии в десетина столични списания и вестници.

В писмо на проф. Александър Вълканов, директор на Черноморската биологична станция (дн. Варненски аквариум), от 7 юни 1947 г. до акад. Иван Буреш пише: "Николай Боев е един от най-добрите и надеждни студенти-естественици, които са преминавали през последните двадесет и пет години през нашия университет. По отношение на неговата начетеност и познания аз го поставям без колебание в редицата на малкото наши вече напълно оформени зоолози. Основа, която го издига особено високо в моите очи и представлява сигурен залог за неговия успех и напредък в науката, е кристално чистата му любов към зоологията, на която се е посветил, любов, която го държи много високо над користното отношение към науката." На 14 юни 1947 г. проф. Георги Паспалев пише до акад. Буреш: "... през време на студентството си г-н Боев проявяваше особен интерес към зоологичните дисциплини изобщо, но особено към тия, засягащи гръбначните животни и по-специално към птиците и бозайниците. Благодарение на този интерес той има върху тия гръбначни животни доста познания, които надхвърлят даваното в университетския курс. Същият ... работи дипломна работа при мене и аз съм с впечатление, че той има нужните данни за научни занимания." Висока оценка на качествата на Николай като учен-зоолог дава и Константин Христович, притежател и редактор на списанието "Природа" в писмо до д-р Буреш от 15 юни 1947 г.: "... познавам го като младеж с добри познания върху птичата и бозайна фауна в нашата страна. Публикуваните от него в редактираното от мене списание "Природа" статии намирам за много сполучливи и заслужаващи насърчение.". С такива отзиви от най-изтъкнатите наши естественици Боев постъпва на работа в Българска академия на науките. Двадесет години по-късно, при хабилитирането му, акад. Ив. Буреш отбелязва: "Той правеше (като студент – б.а.) впечатление на всички свои професори с голямата си любознателност. Много отрано той влезе във връзка с тогавашния директор на зоологическата градина, орнитолога Павел Патев. Двамата се проявиха в онова време преди 30 години (т.е. 15 – 16 годишен) като единствените орнитолози в България. ... Николай Боев много отрано се прояви като голям книголюбител. Той редовно посещаваше всички книжарници и антиквариати в София и с последната си стотинка (като студент, а и по-късно) купуваше природонаучни книги. В кратко време той натрупа голяма и богата библиотека, която днес може да прави чест на всеки орнитологически институт." Тази негова любов към книгите е вдъхната от баща му и дядо му Никола Боев, за които тя е била и професия. В началото на юни 1947 г. Н. Боев завършва семестриално следването си във Физико-математическия факултет. Дипломира през 1949 г.

Професионален път
На 1 август 1947 г. е назначен като лаборант в Природонаучния музей при БАН (дн. Национален природонаучен музей), а от 1 юни 1949 г. – като асистент в Зоологическата градина при БАН. От 1 март 1950 г. е назначен за младши научен сътрудник в Зоологическия нститут с музей при БАН (преди това – Институт по зоология, след преминаването на Зоологическата градина от БАН към Столичната община), но след 31 ноември 1951 г. пак е прехвърлен на работа в зоопарка. През 1963 г. е върнат отново на работа в Зоологическия институт с музей, където работи до 1969 г. Цялата му трудова дейност преминава в учрежденията на Българската академия на науките. Като лаборант в музея, Николай Боев подрежда разпилените и повредени от бомбардировките колекции от птици, яйца и гнезда и помага на д-р Нено Атанасов за подреждането на колекциите от бозайници, на д-р Буреш – за влечугите и земноводните, а на доц. Пенчо Дренски – за рибите. Оказва съдействие в и набавянето на необходимата научна специализирана литература по зоология за библиотеката на института.

От първите години на научната си кариера, Н. Боев е ревностен защитник на природата. Той става член на Съюза за защита на родната природа и изготвя неговата емблема – профилното изображение на брадат лешояд, впоследствие превърнало се в символ на българската природозащита. От май 1951 г. Н. Боев е секретар на Съюза за защита на природата с председател д-р Кирил Ризов. По-късно членува и в Комисията по защита на природата при БАН до трансформирането ѝ през 1967 г. По негова инициатива пред Британското посолство у нас в Природонаучния музей се организира изложбата "Защита на природата в Англия", а в Лондон – на "Защита на природата в България". Участва в работата на комисията по изработването на текста на Закона за защита на природата. Боев е инициатор за издаването и съавтор на първия в България учебник по защита на природата. Активната популяризация на природозащитните идеи го превръща в един от най-изтъкнатите наши популяризатори на природонаучни и природозащитни знания.

Дванадесетгодишната му дейност като асистент и научен сътрудник в Софийската зоологическа градина при БАН (дн. Софийски зоопарк) се изразява в подобряване на съществуващите помещения за животните и разширяване на експозиционната площ с нови постройки, определяне на редовни сезонни дажби за животните, както и на специални дажби за млади животни, бременни женски и др., набавяне на нови експонати от местната фауна, комплектуване на сбирките от живи ескпонати чрез набавяне на нови екземпляри чрез обмен с чуждестранни зоопаркове и зоофирми, ежедневен контрол на състоянието на експонатите от поверения му сектор в зоопарка – птиците, изработване на научно верни текстове с информация за животните, съставяне на пътеводител за зоопарка, научноизследователска дейност, проучване на специалната литература за изграждането на зоопарковете или отделни сектори и клетки за конкретни видове. Цялата тази дейност, проведена с вещината на компетентен специалист, е от изключително значение за твърде обеднялата на видове и екземпляри след войната зоологическа градина. Н. Боев е консултант и на проектите и съпроектант на зоопарковете в Хасково, Димитровград, Благоевград, Габрово, Варна и др., както и на новия Софийски зоопарк, от който е реализиран само първият етап. Най-интересните епизоди от "зоопарковия" период в научното поприще на Н. Боев намират отражение в книгата му "И птиците обичат"

Като млад специалист в БАН той завежда Българската орнитоцлогична централа, отговаря за стопанисването на Мандренското езеро, дн. язовир Мандра, активно сътрудничи на тогавашната Димитровска пионерска организация "Септемврийче", изготвя текстове за всички клетки на животните, води кореспондеция с ловните дружинки и с чуждестранните зоопаркове по различни въпроси на отглеждането на животните и др. През тези години използва възможността да се запознае с дейността и устройството на зоологическите градини в бившите източен и западен Берлин, Дрезден, Лайпциг, Хелзинки, Стокхолм, Лондон, Амстердам, Прага, Братислава, Острава, Варшава, Вроцлав, Букурещ, Москва, Санкт-Петербург, Киев, Одеса, Харков, двата зоопарка в Париж, херпетологичната къща в Неапол и др. Всичко това го прави един от най-добрите специалисти и по зоопарково дело в България. През 1951 – 1953 г. като служител в зоопарка, Боев работи върху 3 научноизследователски теми: "Очерк върху птиците в Софийско", "Състояние на блатния водоплаващ дивеч и мерки за неговото увеличение" и "Птиците на Добруджа".

Най-плодотворният период за Николай Боев като полеви орнитолог е десетилетието 1955 – 1965 г., когато той съобщава 4 нови вида (тук се нямат предвид 30-те вида птици, съобщени през 1944 г. на П. Патев) за българската орнититофауна – среден корморан (Phalacrocorax aristotelis), трипръста чайка (Rissa tridactyla), дългоклюна чайка (Larus gennei) и тънкоклюн листоног (Phalaropus lobatus), проучва с колеги състава и разпространението на птиците в Тракия, публикува много нови данни за редица слабо известни в страната видове и отпечатва най-пълната за времето си фауна на птиците в България, съдържаща много данни от лични наблюдения и илюстрирана от него с черно-бели щрихови рисунки . Н. Боев воюва за прекратяване на "пушечния" етап от развитието на българската орнитология. За установяване на наблюденията той предпочита използването на зрителна тръба и бинокъл, вместо отстрелване на "доказателствения" екземпляр, така както е от десетилетия в по-надпредналите страни в Европа.

През 1962 г. Н. Боев води група английски орнитолози по време на 45-дневното им пътуване из страната за извършване на орнитологични наблюдения. Сред тях са бележитият орнитолог Guy Mountfort, световноизвестният фотограф-анималист Eric Hosking и др. Впоследствие д-р Маунфорт написва книга, по-голямата част от която разкрива великолепието на природата на България. Оценен високо от английските орнитолози, Н. Боев е поканен лично от секретаря на Международния орнитологичен конгрес в Оксфорд, Nikolaas Tinbergen (Нобелов лауреат, 1973) за участие в него. Укрепването на научните контакти между българските и британските орнитолози след 15-ина години изпитва нов тласък с определянето на Н. Боев от Държавен комитет за наука и технически прогрес при Министерския съвет за научен ръководител на английския орнитолог John Allan Roberts, който в продължение на няколко години публикува редица свои трудове с интересни и нови данни за българската орнитофауна в реномирани британски издания.

През 1963 г. по инициатива на проф. Георги Паспалев, Боев е поканен да чете леции по зоогеография и специален курс по орнитология в Биологическия факултет в Софийския университет, което прави в продължение на 5 години.

По покана на Централния научноизследователски институт по горско стопанство Н. Боев изследва орнитофауната в района на язовир Искър и в полезащитни горски пояси в Южна Добруджа. Член е на Висшия ловен съвет при Министерството на горите и с участието си допринася много за утвърждаването на правилно отношение към стопанисването и опазването на дивечовите ресурси на страната. Заслуга на Н. Боев е прекратяването на анахроничната срамна практика организираните ловци в Български ловно-рибарски съюз (дн. Съюз на ловците и риболовците в България) да се отчитат за дейността си по унищожаването на вреден дивеч с представянето на отрязани крака от орли, ястреби, соколи, премахването на гюметата като начин за ловуване на блатен дивеч и др. Най-активно Боев участва в изготвянето на новия Закон за защита на природата и Правилника за приложението му.

Пак по това време (1963 – 1965 г.) като уредник в музея той изготвя план за "постепенна модерна реорганизация", въвежда нов тип на експониране в биогрупи и "малки макетни диорами", провежда инвентаризация и значително допринася за обогатяването на музейните колекции с нови препарати. Продължава работата си по съставянето на "Български зоологичен речник", който смятал да послужи за изготвянето на българската зоологична номенклатура, каквато и до днес все още няма изградена. Към 1968 г. той вече е събрал над 2000 български народни названия на птиците (средно по 6 имена за всеки вид) и още много други за останалите животни. По подобие на труда си за изчезването на бобъра (Castor fiber) у нас . Към 1968 г. Боев е почти готов с публикуването на работите си за тура (Bos primigenius) и елен лопатар (Cervus dama). Те обаче остават непубликувани.

На 13 декември 1967 г. след участие в конкурс, от 1970 г. е Н. Боев е назначен като старши научен сътрудник в Комисията по защита на природата при БАН с председател проф. Георги Паспалев, където е неин научен секретар. Впоследствие става ръководител на Секция "Защитени природни територии и защитен генетичен фонд". Там дейността му е свързана главно с проучването и организирането на защитени природни територии (национални паркове, резервати, защитени местности и природни паметници), както и с конкретни мерки за опазване на редките и застрашени видове.

През 1976 – 1977 г. Николай Боев работи по няколко научни теми върху аклиматизацията на диви бозайници и птици, изчезването на редки видове, методи за проучване и опазване на резервати и други защитени територии и обекти, защитени видове животни, природозащитни изследвания на Разградски окръг и др. По това време е и член на научните съвети при издателствата "Наука и изкуство", "Земиздат" и "Варна", в които успява да предложи за отпечатване у нас на десетки полезни и интересни популярни книги с природонаучна и природозащитна тематика на известни чуждестранни автори като Жан Дорст, Бернхард Гжимек, Джой Адамсън, Максим Зверев, Джералд Даръл, Борис Стрелников, Никола Тинберген, Юлиус Комарек, Рейчъл Карлсън, Джейн Гудол, Е. П. Спангенберг и др.

Боев мечтае и за изграждането у нас на един Институт по екология и защита на природата, каквито вече имало в редица източно-европейски страни. Той смята, че НКЦОВОС би могъл да прерасне в едно такова научно учреждение към Българската академия на науките. Изготвя концепция, и подробни планове, с които предлага изграждането му пред ръководството на БАН и висши държавни институции. За жалост тази негова идея, макар и за кратко, се реализира едва след смъртта му. Впоследствие Институтът по екология при БАН е трансформиран в Централна лаборатория по обща екология при БАН, а в 2010 г. става част от Институт по биоразнообразие и екосистемни изследвания на БАН.

В периода 1973 – 1976 г. Н. Боев отново е поканен за преподавател в Биологическия факултет на Софийския университет – този път по "Основи на защитата на природната среда". През 1979 – 1981 г. той често е командирован в гр. Котел, където работи като научен консултант по изграждането на експозицията на новия Природонаучен музей, открит за посещеия на 5 октомври 1981 г. Заедно с това работи (1979 – 1980 г.) и по своята част за птиците от неколкократно отлаганата за печат "Червена книга на Н Р България" В нея той е автор на статиите за 16 вида птици.

В началото на 80-те години Н. Боев вече е събрал огромно количество данни и по три други любими свои теми, по които е възнамерявал скоро да публикува обобщените си резултати: "Топонимиката като относителен биогеографски индикатор и в помощ на природозащитата", "Към историята на природозащитата у нас през Първата и Втората българска държава и Отоманското иго" и "Материали за проучването на съществуването на тура у нас". През 1981 г. той планира издаването на книгите си "Аклиматизация на дивите бозайници и птици" и "Животни и топонимия". Усилено работи и върху "Биографии на 100 животни", замислена в два тома (за използваните от човека видове животни). Предвижда да издаде второ поправено и допълнено издание на частта за птиците от "Фауна на България – кратък определител", както и на популярната му книга "Грижи за поколението у животните" в 2 части. Той не успява да отпечата тези интересни свои трудове. Пенсионира се през 1984 г. Умира на 12 ноември следващата година, работейки над коректурите на последната си книга – за декоративните птици.

Николай Боев е бил делегат от България на десетки научни конгреси, сипозиуми, конференции и съвещания в чужбина. Като орнитолог е бил делегат на международните орнитологични конгреси в Хелзинки, Оксфорд и Москва, на орнитологичните конференции в Прага и Ленинград, на конгреса по ловознание в Москва и мн. др.

По време на една командировка във Франция, в Големия салон на Природонаучния музей в Париж Боев изнася доклад пред повече от 300 естественици за опазването на животинския свят в България. Така запознава френската научна общественост със състоянието на българската природозащита, от която дотогава на запад са нямали и представа.

Популяризиране на природозащитната идея
Научното творчество на Боев се състои в 50 научни труда, 4 учебника, 29 научно-популярни книги и около 650 научно-популярни и публицистични статии. Сред по-значимите му трудове са тези за бобъра, средния корморан, степния пор, гугутката , за черния щъркел, птиците на Тракия, частта за птиците във "Фауна на България – кратък определител" и др. Някои от научно-популярните му книги са преведени на чужди езици и издадени в чужбина – три издания в Германия, по едно – в Турция и Чехословакия. Популярните му статии са предимно на зоологична и природозащитна тематика и някои са преведени и издадени в чужбина на английски, френски, италиански, немски, чешки и други езици. От книгите си, самият Н. Боев определя като по-интересни "Прелета на птиците" , "Нашите полезни птици" , "Полска практика по зоология".

Книгата му за полската яребица  е отпечатана в съкратен превод на руски език в списанието "Охота и охотничье хозяйство" – едно от най-известните ловни списания, и на хърватски език в списанието "Lovac". Книгата му "Детско букварче за растения и животни" претърпява 5 издания, а преведената от него и Ат. Георгиев книга "Земя отвъд океана" – 3 издания. Най-значимата от популярните му книги обаче, "Декоративни птици", излиза посмъртно, една година след кончината му. Три години по-късно (1988) е създадено (по негова идея) Българско дружество за защита на птиците. Дванадесет години по-късно излизат и две основни статии за вълка в България – една от последните теми, по които работи . За научно-популярните си статии той отбелязва: "... за тях нито водя точен списък, нито имам от всички на разположение, а нерядко някои от тях не носят и моето име, а друго или са неподписани." Според него до 1967 г. броят им надхвърля 350.

Н. Боев е инициатор за издаването на "Червена книга на Република България" в два тома, неин съавтор и един от главните ѝ редактори. Боев мечтае и за издаването на една модерна тритомна "Птиците на България" от поредицата "Фауна на България (монография)" и въпреки че към 1967 г. е направил доста за подготвянето му, това издание се появява след повече от две десетилетия. Макар и доста закъснял, първият том се появява през 1990 г. Автор е и на повече от 120 статии за птиците в 5-томната "Кратка българска енциклопедия" (1963 – 1969) и на статиите за животинския свят в "Енциклопедия А – Я" (1974), издадени от БАН, част от които са илюстрирани лично от него.

Като един от най-изявените популяризатори на природонаучни знания, Николай Боев е бил член на редакционните колегии на списанията: "Лов и риболов" (от 1955 г.), "Бодра смяна", "Защита на природата" (от създаването му през 1970 г.), "Наука и техника за младежта" (от 1967 г.), "Турист", "Курорти", "Наша Родина", "Природа и знание", "Космос", на туристическия вестник "Ехо" и вестниците "Вечерни новини" и "Септемврийче". Той е консултант при Държавната кинематография – Студия за научно-популярни филми и е член и на Висшия ловен съвет при Министерството на горите и горската промишленост. Автор е и на 10 сценария за научно-популярни филми с природозащитна и зоологична тематика. С научната си дейност и огромните му приноси в областта на популяризацията на защитата на природата Николай Боев е смятан за основоположника на съвременната природозащита в България. За него авторитетното британско орнитологично списание "The Ibis" пише: "[Боев] е бащата на съвременната орнитология и природозащита в България. … Беше привилегия да го познаваш.".  8 май (рождената дата на Николай Боев) е включен в природозащитния "Календар" на България.

Бележки

Източници

Български зоолози
Български орнитолози
Български природозащитници
Родени в Айтос
Починали в София

redpajama_set_name: RedPajamaWikipedia


These tournaments are using SpeechWire during the 2018-2019 season and meet the criteria above. A few links may appear under 'More information'. 'Info' will let you learn more about a tournament, 'Register' will send you to tournament registration (in your SpeechWire account), and 'Results' will show you the tournament results.


Please take a moment prior to your scheduled appointment to print our patient forms. We ask that you complete the forms and bring them with you to your appointment. If you have any questions while reviewing this information please feel free to contact our office.
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The Holidays are one of the best times to make pictures. (Check out Bryan's and Lauri's articles for some terrific ideas.) And it's a great time to photograph kids–if you haven't discovered it, yet, a simple truth is that the Holidays are better with kids. Here are two simple tips to help you make better pictures with kids, and you should do these everytime you make pictures with children.
Your pictures will be much more engaging if you get the camera on the same level with kids, at the same height as their perspective. In fact, you can affect the feeling of your photos by photographing from the height of different kids. My six-year-old is taller than her little brother but shorter than her cousins and in pictures with the camera at her height, you get a feel for what the world is like from her perspective.
Pointing downward, the background is just the ground.
With the camera on her level, the background is much deeper and out of focus.
Another great advantage of getting low is that you get a much deeper background rather than having only the ground right behind your subject as the backdrop. Putting the background farther away makes it more out of focus and it feels like a more "professional" picture.
Kids love to play and playing with them is much more important than photographing them. One of the best reasons to make pictures is to capture memories to share and relive later, but if you're so busy making pictures that you don't make memories then your pictures may be lovely but the memories will be empty. Or worse, your kids will remember that you didn't play because you were making pictures and their memories will be full of resentment. So, put the camera down and play! Make a picture while you can but remember the important stuff.


With the same rules as the original Mapominoes - much the same principle as double six dominoes - in Mapominoes UK you build a map of Britain by connecting counties sharing a common border. Race to be the first player to play all your county cards on the table and win the game. Block access to the Scottish Borders by holding on to Northumberland or Cumbria or open up Wales by playing Powys. Take a boat through the Irish Sea to Northern Ireland or travel by train through Surrey to Hampshire or Berkshire. Contents: 54 county cards, 12 transit cards, sand timer, rules.