Purpose: To investigate the potential protective effects of total flavonoids from Flos Puerariae (TFF) on retinal neural cells in diabetic mice.
Methods: C57BL/6J mice were intraperitoneally injected with streptozotocin to generate type I diabetes in a murine model, as indicated by blood glucose levels ≥11.1 mmol/l. TFF was administered intragastrically at a dose of 50, 100, or 200 mg/kg/day. After 10 weeks of administration, the mice were euthanized, and the eyes were dissected. Retinal histology was examined, and the thickness of the retina was measured. Ultrastructural changes in the retinal ganglion cells and capillary basement membrane were observed with electron microscopy. Apoptosis of retinal neural cells was determined with the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling assay. Bax and Bcl-2 expression in the retinal tissues was determined with immunohistochemical staining and western blotting.
Results: Compared with the diabetic mice, the blood glucose level decreased (p<0.01) and the bodyweight increased (p<0.05) in the 100 and 200 mg/kg TFF-treated groups. The thickness of the retina significantly increased (p<0.01), and the retinal capillary basement membrane (BM) thickness was reduced in the 100 and 200 mg/kg TFF-treated diabetic mice (DM). The 100 and 200 mg/kg TFF treatments also attenuated the diabetes-induced apoptosis of retinal neural cells. Consistent with these effects, TFF treatment decreased the Bax expression level and, concurrently, increased the ratio of Bcl-2 to Bax.
Conclusions: TFF attenuated diabetes-induced apoptosis in retinal neurons by inhibiting Bax expression and increasing the ratio of Bcl-2 to Bax, which suggests that TFF might prevent retinal neuronal damage in diabetes mellitus.
Diabetic retinopathy (DR) is the most common complication of diabetes mellitus and one of the major causes of vision loss across the globe. Almost all patients with diabetes suffer different degrees of retinopathy after 20 years' duration of diabetes . It has long been believed that DR is a microvascular disease . In recent years, an increasing body of evidence has suggested that neuronal cell death of the retina is a critical component of DR [3-7]. With microvascular lesions, neurodegeneration might occur in the early stage of diabetic retinopathy, preceding retinal vascular complications in humans and experimental animals [8,9]. Abu El-Asrar et al. reported that ganglion cells in diabetic retinas express several proapoptotic molecules, such as caspase-3, Fas, and Bax, suggesting that these cells are the most vulnerable population in diabetic retinopathy . Another study showed that neurotoxicity causes permanent impairment of visual function due to cell death of the inner retinal and ganglion cells . Intervention in the apoptosis of ganglion cells may allow us to therapeutically delay or ameliorate neural cell loss in retinal neurodegenerative conditions related to diabetes.
Traditional Chinese medicine has been widely used for centuries and can play a unique therapeutic role in treating many human diseases. Flos Puerariae, a well known Chinese medicine compound, is the dry bud of Pueraria lobata (a plant in the genus Pueraria in the pea family Fabaceae, subfamily Faboideae). Seven isoflavones have been identified in the Flos Puerariae extract, including four isoflavone glycosides (tectoridin, tectorigenin7-oxyloglucoside, 6-hydroxy-genistein-6,7-diglucoside, and glycitin) and three aglycones (tectorigenin, glycitein, and genistein). Three saponins (soyasaponin I, kaikasaponin III, and kakkasaponin I) have been identified in the extract [11-13]. The extract of Flos Puerariae has been reported to have a wide range of pharmacological effects, including prevention of the effects of excessive alcohol ingestion (hangovers) , antioxidant action , a liver protective effect , an estrogenic-like effect , hypolipidemic and hypoglycemic effects [18,19], and an antiapoptotic effect [20,21]. Considering the close correlation between DR and the apoptosis of retinal neural cells, total flavonoids from Flos Puerariae (TFF) may improve DR by hypoglycemia and inhibiting neuronal damage.
To test this hypothesis, we examined whether TFF has a neuroprotective effect on retinal neural cells in the streptozotocin (STZ)-induced diabetic mouse model. The regulation of various apoptotic-related genes, such as Bax and Bcl-2, has been used to evaluate apoptotic activity in tissues. The balance of Bax and Bcl-2 ensures a steady-state of the cell, and changes in the Bcl-2/Bax expression ratio determine cell survival or apoptosis [21-23]. In this study, we investigated the effect of oral administration of TFF on hyperglycemia and the morphology of the diabetic mouse retina, as well as the effect on apoptosis of the retinal neurons in relation to the expression of apoptotic-related genes (Bax and Bcl-2).
In this study, total flavonoids were extracted from Flos Puerariae using an ultrasonic-assisted extraction method as previously reported . Briefly, Flos Puerariae was extracted in 40% methanol and broken down by ultrasound for 4 h. Petroleum ether was blended with the brown viscous extract, and the extract was separated into two layers, a petroleum ether layer (to remove fat-soluble impurities) and a brown water layer. The brown water layer was collected and concentrated with anhydrous ethanol. The dark brown sediment (predominantly polysaccharide) was precipitated. The supernatant ethanol solution was concentrated with anhydrous ethanol. The remaining solution comprised the total flavonoids extracted from Flos Puerariae.
Fifty male C57BL/6J mice, weighing 22±2.0 g, were purchased from the Wuhan University Center for Animal Experimentation. Animals were housed in a specific pathogen-free animal house according to the guidelines established by the Association for Research in Vision and Ophthalmology's Statement for the Use of Animals in Ophthalmic and Vision Research. The animals were fed by adaptive feeding (with free food and water) for 1 week. They were housed with a 12 h:12 h light-dark cycle at 23–25 °C and 55%–60% humidity.
Forty C57BL/6J mice were intraperitoneally injected with STZ (Sigma Aldrich, St. Louis, MO) to generate the type I diabetic model, as described previously [25,26] with modifications. We induced diabetes in mice with multiple intraperitoneal injections of STZ dissolved in sodium citrate buffer (pH 4.5) at a low-dose level of 50 mg/kg bodyweight daily for 5 consecutive days. Ten age-matched mice were given the same volume of sodium citrate instead of STZ and served as controls. A sample of whole blood was drawn from the tail vein of each mouse by cutting the tail, and blood glucose was measured with a glucometer (Sinocare Inc., Changsha, China) at 2 and 4 weeks post-injection. After 4 weeks of exposure to STZ, 36 mice with blood glucose levels ≥11.1 mmol/l were considered diabetic. They were randomly divided into four groups (n=9, each group): the diabetic group (DM), the 50 mg/kg/day TFF-treated group (DM+TFF I), the 100 mg/kg/day TFF-treated group (DM+TFF II), and the 200 mg/kg/day TFF-treated group (DM+TFF III). The mice in the drug intervention groups had intragastric TFF administration daily. The mice in the DM group did not receive treatment. The mice were weighed weekly, and blood glucose was measured at the end of the course of treatment. After 10 weeks of TFF administration, the mice were euthanized by intraperitoneal injetion of 1% pentobarbital (50 mg/kg), and the eyes were dissected. Four eyes from four mice in each group were immediately fixed in 4% paraformaldehyde solution for histopathological examination. The remaining retinas were carefully dissected under a stereomicroscope. Three retinas from three individuals in each group were immediately placed in a 2.5% glutaraldehyde solution for ultrastructural analysis. The remaining retinas were frozen immediately in liquid nitrogen and stored at −80 °C for western blotting analysis.
Four paraformaldehyde-fixed eye tissues from each group were embedded in paraffin and sectioned at 5 µm thickness each for hematoxylin and eosin (H&E) staining, terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) analysis, and immunohistochemical staining. All color micrographs were collected using a microscope and digital camera (Axioplan 2 imaging; Carl Zeiss, Inc., Shanghai, China; camera equipped with Spot Software ver. AxioVision Rel 4.8). The morphology of the retina, including measurements of the thickness of the total retina, inner nuclear layer (INL), and outer nuclear layer (ONL), was determined with Spot Software ver. AxioVision Rel 4.8. The thickness was measured on the magnified images (×400) of the retinal equator at six points: three on either side of the retinal equator, located approximately 500 to 600 μm apart from the ora serrata. The mean thickness value of the four eyes was recorded as the representative value for each group. The results of the five groups were analyzed with ANOVA (ANOVA).
The TUNEL assay was performed using the TUNEL Apoptosis Assay Kit (Roche Applied Science, Nutley, NJ) according to the manufacturer's instructions. Three sections from each group were selected for the TUNEL assay. The dewaxed sections were treated with 20 μg/ml proteinase K for 15 min, incubated with equilibration buffer for 15 s, and incubated with terminal deoxynucleotidyl transferase (TdT) enzyme at 37 °C for 1 h. Antibody blocking proceeded for 5 min, and peroxidase (POD; conjugated with horseradish peroxidase) was incubated with the sections. The sections were incubated with substrate 3,3′-diaminobenzidine for 8 min.
Three visual fields (×400) under microscope (Axioplan 2 imaging) per section were randomly chosen to count the number of TUNEL-positive cell nuclei (apoptotic nuclei) and total cell nuclei in the ganglion cell layer (GCL) and the INL, respectively. The percentage of TUNEL-positive cells, termed the apoptotic index (AI), was calculated using the following formula: AI=apoptotic nuclei/total nuclei ×100%. The mean value of the eyes from four mice was recorded as the representative value for each group. The results of the five groups were statistically analyzed with ANOVA.
To examine Bax and Bcl-2 protein expression in the retinal tissue, immunohistochemical staining was performed on paraffin sections as previously reported . Three sections of each paraffin-embedded eye tissue from each group underwent immunohistochemical staining. Tissue sections were incubated with polyclonal primary antibodies against Bax or Bcl-2 (Santa Cruz Biotechnologies, Santa Cruz, CA) overnight at 4 °C. After being washed with PBS (phosphate buffer saline), the sections were incubated with biotinylated horse anti-mouse immunoglobulin G for 30 min and then incubated with the avidin-biotin-peroxidase complex using an ABC kit. The reaction was visualized by color development with 3, 3′-diaminobenzidine tetrahydrochloride. All sections were counterstained with hematoxylin. Images from the immunohistochemical studies of Bax and Bcl-2 protein expression (×400) were photographed with a microscope and digital camera (Axioplan 2 imaging; the camera was equipped with Spot Software ver. AxioVision Rel 4.8).
Three pieces of retina approximately 2 mm × 3 mm in size were isolated from each eyecup, approximately 2 mm from the optic disc, following the protocol described previously . Briefly, retina tissues were immediately prefixed in 2.5% glutaraldehyde for 2 h and then fixed in 1% osmium tetroxide. Sequentially, the tissues were dehydrated and embedded in Epon 812. The tissues were sectioned at a thickness of 50–60 nm using an ultramicrotome and double-stained with uranyl acetate and lead citrate. The ultrastructure of the retina tissues was observed using a transmission electron microscope (TEM H-600, Hitachi, Tokyo, Japan).
Western blot tests were performed as described previously . The retinas in each group were pooled and homogenized in ice-cold lysis buffer (20 mM Tris, pH 7.5, 150 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1% Triton X-100, 2.5 mM sodium pyrophosphate, 1 mM β-glycerolphosphate, 1 mM Na3VO4, 1 μg/ml aprotinin leupeptin and pepstatin, and1 mM phenylmethylsulfonyl fluoride) and centrifuged at 12,890 ×g for 15 min at 4 °C. The supernatant was collected, and the protein concentration was measured using the bicinchoninic acid protein assay (Beyotime, Jiangsu, China). An equal amount of protein for each sample (60 μg/lane) was separated on 10% sodium dodecyl sulfate–polyacrylamide gel and transferred to a polyvinylidene fluoride membrane. The membranes were blocked in 5% skim milk for 2 h at room temperature before incubation with an antibody against Bax, Bcl-2 (Cell Signaling Technology, Boston, MA), or β-actin (Santa Cruz Biotechnology, Dallas, TX) overnight at 4 °C. The membranes were incubated with horseradish peroxidase-conjugated goat anti-mouse immunoglobulin G (GE Healthcare, Buckinghamshire, UK) for 2 h, and blots were developed using an enhanced chemiluminescence kit (Pierce Biosciences, Rockford, IL). β-actin expression was used as an internal loading control to standardize the amount of loaded protein. Experiments were repeated five times. The intensity of the protein band was semiquantitatively measured with image analysis software (Image-Pro Plus 6.0, Media Cybernetics, Bethesda, MD).
All data were reported as the mean±standard deviation. One-way ANOVA and Student–Newman–Keuls tests were performed to compare the means of the groups using statistical software (SPSS Statistics 17.0, New York, NY). A value of p<0.05 was considered statistically significant.
After the full course of exposure to STZ, as shown in Table 1, the blood glucose levels of the diabetic mice significantly increased compared to those of the control mice (20.12±6.66 mmol/L versus 6.34±0.89 mmol/l, p<0.01). Ten-week 100 and 200 mg/kg TFF treatments decreased the blood glucose levels of the diabetic mice compared with the untreated diabetic mice (12.39±5.25 and 11.17±4.39 mmol/l versus 20.12±6.66 mmol/l, p<0.01).
The line graph (Figure 1) shows the change in bodyweight for each group at each week of treatment. The initial bodyweight in each group was similar. The bodyweight of the control mice increased steadily over the 14-week course. At 4 weeks after the onset of the STZ treatment, the bodyweight of the diabetic model mice began to decrease. Subsequently, the bodyweight of the diabetic mice decreased 5.11% by 14 weeks. The bodyweight of the 50 and 100 mg/kg TFF-treated mice remained relatively stable and increased 1.89% and 2.67%, respectively, by 14 weeks. The bodyweight of the 200 mg/kg TFF-treated mice increased 8.22% by 14 weeks.
A systematic morphology examination of the H&E-stained retinal sections (Figure 2) showed that the overall thickness of the retina in the diabetic mice was reduced by 19.58% compared with the control group (131.45±17.28 versus 163.45±13.58 μm, p<0.01). The thickness measurements of the INL and the ONL decreased by 18.17% and 18.99%, respectively, compared with the control group. In the diabetic mice treated with 50, 100, and 200 mg/kg TFF group, the retina appeared markedly more normal (Figure 2A), and the thickness of the overall retina increased compared with that of the diabetic group (145.46±17.05, 151.35±6.58 and 159.85±24.26 versus 131.45±17.28 μm, p<0.01). The thickness of the INL in the diabetic mice treated with 100 and 200 mg/kg TFF was significantly thicker than that in the diabetic group (21.84±1.99 and 22.18±3.22 versus 18.21±3.60 μm, p<0.01). The thickness of the ONL in the diabetic mice treated with 50, 100, and 200 mg/kg TFF was significantly greater than that in the diabetic group (37.21±4.49, 38.34±3.04 and 39.48±8.17 versus 33.95±4.92 μm, respectively, p<0.05).
Retinal ultrathin sections were examined systematically with a transmission electron microscope to determine whether TFF affects apoptosis of the retinal ganglion cells (RGCs) and thickening of the retinal capillary basement movement (BM). The RGCs in the normal control mice had uniformly normal-appearing nuclei and mitochondria (Figure 3A,F), while vacuolation, chromatin margination, chromatin condensation, and swollen mitochondria were detected in the RGCs of the diabetic mice (Figure 3B,G). In the 50 mg/kg TFF-treated group, chromatin condensation and swollen mitochondria were detected (Figure 3C,H). The RGCs in the diabetic mice treated with 100 and 200 mg/kg TFF did not show obvious vacuolation, chromatin margination, chromatin condensation, or swollen mitochondria (Figure 3D,I,E,J).
An electron microscopy analysis of the retinal capillary (Figure 4) showed that the BM thickness of the retinal capillary was significantly increased in the diabetic mice compared to the normal control mice (0.16±0.05 versus 0.09±0.02 μm, p<0.05). After treatment with TFF, a significant decrease in retinal capillary BM thickness was observed in the retinas of diabetic mice compared to those of the diabetic model mice. The BM thickness of the retinal capillary in the 100 and 200 mg/kg TFF-treated group was significantly decreased compared with that of the diabetic model group (0.12±0.06 and 0.10±0.02 versus 0.16±0.05 μm, p<0.05).
An examination of the retinas of diabetic mice showed more TUNEL-positive cells in the GCL and the INL than in the control mice (Figure 5A). The presence of TUNEL-positive cells was not detected in the outer nuclear layer. There were fewer TUNEL-positive cells in the GCL and the INL in the TFF-administered groups than in the diabetic model group, specifically in the 200 mg/kg TFF-treated group (Figure 5A). A quantitative analysis of the TUNEL-positive cells in the GCL and the INL of the retinas (Figure 5B,C) showed that the AI (apoptotic index, apoptotic nuclei/total nuclei ×100%) in the GCL and the INL of the diabetic mice increased eightfold and sevenfold, respectively, compared to that in the control mice. The 100 and 200 mg/kg TFF treatments significantly reduced the AI in the GCL compared with the diabetic group (51.18% and 27.16% versus 73.50%, p<0.01). The 50 mg/kg TFF treatment slightly reduced the AI in the INL compared with the diabetic group (44.95% versus 53.28%, p<0.05). The 100 and 200 mg/kg TFF treatments significantly reduced the AI in the INL compared with the diabetic group (35.28% and 13.35% versus 53.28%, p<0.01).
Bax and Bcl-2 expression, which is related to apoptosis, was observed using immunohistochemistry staining and western blotting (Figure 6). Immunohistochemistry staining showed that the Bax and Bcl-2 positive signal was localized in the cytosol of cells in the GCL and the INL. The immunohistochemistry and western blot analyses revealed that Bax expression significantly increased and Bcl-2 expression relatively decreased in the diabetic group compared with the control group. However, the 100 and 200 mg/kg TFF treatments decreased Bax expression compared with that of the diabetic group. The relative intensities of Bcl-2 (as normalized to the signal of β-actin) of each group showed no significant differences. As shown in Figure 6E, the expression ratio of Bcl-2 to Bax was significantly decreased in the diabetic mice retinas than in the control group (0.61±0.31 versus 2.98±0.95, p<0.01). The 100 and 200 mg/kg TFF treatments enhanced the ratio of Bcl-2 to Bax compared to the ratio in the diabetic group (1.97±0.39 and 2.37±1.01 versus 0.61±0.31, p<0.01). The ratio of Bcl-2 to Bax (Bcl-2/Bax) demonstrated the ability of TFF to inhibit apoptosis of the retina by upregulating Bcl-2 expression and downregulating Bax expression.
This study investigated the protective effects of TFF in preventing retinal neurocyte damage in STZ-induced diabetic mice. TFF reduced the blood glucose levels of diabetic mice and decreased diabetic-induced bodyweight loss in our study. We found that TFF markedly reduced retinal neuron injury related to diabetes, as demonstrated by an improvement in histological damage compared to the diabetic groups. We found that TFF prevented apoptosis of the retinal neurons by upregulating Bcl-2 expression and downregulating Bax expression in the diabetic mice, which suggests a possible protective mechanism of TFF in the retina.
Flos Puerariae has been reported to have a wide range of pharmacological effects. The isoflavonoid fraction of Flos Puerariae has been reported to prevent symptoms of excessive alcohol consumption (hangovers)  and to demonstrate antioxidant action . Kakkalide has a liver protective effect , and kakkalide and tectoridin of Puerariae Flosan have an estrogenic effect . Flos Puerariae extract is effective in treating metabolic diseases and displays hypolipidemic and hypoglycemic effects [18,19]. Flavones derived from Radix Puerariae exert inhibitory effects on bodyweight, abdominal fat content, and lipid levels in the liver  in which an increase in visceral fat is responsible for many of metabolic abnormalities, such as impaired glucose tolerance and insulin resistance, associated with abdominal obesity . Kim et al. theorized that genistein may be a potential therapeutic agent for preventing and treating complications associated with diabetes mellitus, such as diabetic cataracts . Aqueous extracts from Flos Puerariae protected against ethanol-induced apoptosis in the human neuroblastoma cell line SK-N-MC [20,21], and tectorigenin, extracted from Flos Puerariae, depressed t-BHP-induced-cellular cytotoxicity that may occur as a result of the inhibition of apoptosis . In the early stage of this study, our colleagues demonstrated that the total flavones of Flos Puerariae (50, 100, and 200 mg/kg; intragastric administration) effectively protect against experimental liver injury induced by alcohol similar to previous research and that the total flavones of Flos Puerariae (50, 100, and 200 mg/kg/day; intragastric administration) could be a potential neuroprotective agent. We hypothesized that neuroprotective effects were achieved, at least in part, by the antioxygen free radical activities of Flos Puerariae, which suggest the potential of the total flavones of Flos Puerariae as an adjuvant therapy to conventional antihyperglycemic regimens for preventing and treating diabetic-associated cognitive decline. That article has not been published, and our study was performed based on those experimental results. We investigated the protective effect of the total flavones of Flos Puerariae on retinal neural cells in diabetic mice rather than fractionate it.
The pathogenesis of DR is complex and is not the consequence of a single mechanism. Neuronal cell death of the retina is a critical component of DR. This study clearly demonstrates the therapeutic potential of TFF in treating diabetic mice. This protective effect of TFF occurred in a dose-dependent manner. We demonstrated that the beneficial effects of TFF against the progression of diabetic retinopathy could be attributed to the hypoglycemic and antiapoptotic properties of TFF.
Studies have suggested that retinal neurodegeneration is a critical component of DR [3-7], which is typically characterized by a reduced number of RGCs, a thinned retina, and an increased number of apoptotic cells. A reduced number of RGCs, a thinned retina, and thinner inner and outer nuclear layers were detected in mice that had been diabetic for 10 weeks . Barber et al.  identified a 22% and 14% decrease in the thickness of the inner plexiform and inner nuclear layers, respectively, of retinal sections from STZ diabetic rats after 7.5 months of STZ-induced diabetes. Oshitari et al.  reported that the number of TUNEL-positive cells in the ganglion cell layer of diabetic retinas was significantly increased compared to that in non-diabetic control rats. In this study, we noticed a significant decrease in retinal thickness and a distinct increase in TUNEL-positive cells in the retinas of diabetic mice. These TUNEL-positive cells were distributed predominantly in the GCL and the INL. The oxygen consumption of the neural cells in the GCL and the INL is abundant; thus, these cells are more sensitive to hypoxia and ischemia . The ganglion cells in diabetic retinas express several proapoptotic molecules, suggesting that these cells are a vulnerable population . These observations are consistent with previous studies. We found that TFF significantly improved morphological changes: The thickness of the entire retina and the inner and outer nuclear layers increased, and we detected decreased apoptosis in the GCL and the INL of diabetic mice. These improvements suggested that TFF can reduce the progression of experimental DR. Our data not only confirmed that apoptosis contributes to retinal neuronal death after the onset of diabetes but also demonstrated that TFF can prevent the apoptosis of retinal neurons. The apoptotic cells showed agglomeration and margination of chromatin , swelling, and vacuolation of mitochondria [36,37]. In the electron microscopy examination, we found obvious vacuolation, swollen mitochondria, margination of chromatin, and accidental apoptotic bodies in the GCL in the diabetic mice retinas, which further indicated that the cells of the ganglion cell layer were damaged via an apoptotic mechanism. There was no obvious change in the GCL of 100 and 200 mg/kg TFF-treated diabetic mice. We assume that TFF prevents the apoptosis of retinal ganglion cells by protecting the mitochondrial ultrastructure and maintaining cell integrity.
Studies on diabetic retinopathy have established a thickened vascular BM as one of the first structural abnormalities related to this condition. Kozak noted capillary BM thickening in the retinas of STZ-induced diabetic rats .Vascular BM serves as a substratum for cell attachment, provides a selective permeability barrier, and regulates cell survival . Due to capillary BM thickening and apoptosis of pericytes, the integrity of the capillary was destroyed, and the blood–retinal barrier was damaged. These changes led to stenosis of the capillary and blood flow change, which promotes retinal ischemia . The exclusive energy source for the retina is glucose. The glucose for retinal nerve activity is transported to the neurons predominantly in two ways: by the retinal artery and vein and by the choriocapillary [41,42]. According to the structural characteristics of the retina, a change in blood vessels may influence the energy supply for retinal neurons. Retinal capillary BM thickening may occur concurrently with neural cell apoptosis or perhaps earlier. This issue should be investigated in future research. In this study, we detected that the retinal capillary BM thickness of diabetic mice significantly increased compared to that of the normal control mice. TFF attenuated the capillary BM thickness of diabetic mice.
The mechanisms of apoptotic processes in retinal neural cells are unclear. Increased cellular apoptosis may be associated with Bax overexpression, which may be responsible, at least in part, for neuronal cell loss in the GCL of diabetic rats . The Bcl-2 family members are involved in regulating activities concerned with the survival and death of neurons through apoptosis [43,44]. Bax, a mitochondrial membrane protein that mediates cell death, was increased in the retina of the diabetic subjects and may contribute to the development of vascular complications in diabetic retinopathy . Several reports indicate that Bax is a critical factor in retinal neuronal cell apoptosis and a target for therapeutic gene strategy to protect damaged retinal neurons [46,47]. Bcl-2, an antiapoptotic member, inhibits cytochrome c release and the activity of proapoptotic members . The intracellular Bcl-2 protein extended cell survival because it specifically blocked apoptotic cell death following various signals . Proapoptotic Bax facilitates cytochrome c release and then activates caspases to induce apoptotic cell death [48,49]. The expression ratio of Bcl-2 to Bax is critical for determining the life span of cells . In this study, a significant increase in Bax protein expression was found in the retina. The expression ratio of Bcl-2 to Bax clearly decreased, as it was implicated in apoptosis of retinal neural cells in previous studies. Administering 100 and 200 mg/kg TFF reduced the expression of the Bax protein and enhanced the ratio of Bcl-2 to Bax. The TFF-induced neuroprotection may have been mediated by normalizing the Bcl-2/Bax level, which attributed to the downregulation of Bax expression and the upregulation of Bcl-2 expression.
We found that TFF protected retinal neurons from undergoing apoptosis in experimental DR. This neuroprotective effect of TFF appeared to be related to the downregulation of Bax and the upregulation of Bcl-2. We concluded that TFF could have potential benefits in preventing the onset and progression of retinopathy in diabetic patients. The protective effects of TFF can be due to multiple beneficial effects on different retinal cells. Further investigation should be undertaken to clarify these beneficial effects.
This work was supported by a grant from the Science and Technology Agency of Hubei Province (Grant No. 2007ABA380). We thank the Hubei Province Key Laboratory on Cardiovascular, Cerebrovascular, and Metabolic Disorders for providing experimental technology and instruments and for giving us much assistance and support.
Tsukahara S, Yamamoto S. Tin-Tin-Win-Shwe, Ahmed S, Kunugita N, Arashidani K, Fujimaki H. Inhalation of low-level formaldehyde increases the Bcl-2/Bax expression ratio in the hippocampus of immunologically sensitized mice. Neuroimmunomodulat. 2006; 13:63-8.


Camp visitors can use a covered volleyball playground, a small football field, bike rental or to walk on one of the three health trails marked with mountain markings.
Within the household there is a souvenir shop with freshly ground coffee, home-made products (honey, jams, brandy, wines, souvenirs on magnets, homemade cheese and more).
A special entrance, accessibility of the terrain, beautiful view and location in greenery, use of other household contents that totally cover about 3 hectares of land, make the stay unforgettable for all who love nature and idyllic countryside. The camp is also "pet-friendly".


Dzulkanean Ali :: New Toy.
Postman Pat delivered them goods at 10:50AM.
Ya boy just got his hands on the new iPhone 7 Plus! Finally managed to pick up the Jet Black from Singtel after what everyone would call a disastrous iPhone purchasing experience despite all their maintenance and "pre-registrations".
I have no regrets buying the Jet Black — even though I know it'll kill me deeply to know there'll be scratches & fingerprints on it.
Anyways, I really can't wait to try out the camera capabilities and see what's in store. There's even 4K video recording which I'm super psyched to get rolling with.
Till the next update with the photos from the 7 Plus!


The week was filled with terrible economic news. For the first time since the Second World War the US had net zero job growth in a month. So much for the promise of green jobs with the hundreds of billion spent by the administration and their department of entropy. In fact the administration is looking mighty dumb this week after Solyndra went bankrupt and this was the poster child of President Obama's new green economy. The backer of Solyndra is a booster for Obama. This means he raised money for Obama's campaign. He was handsomely rewarded by gaining over $550 million of loan guarantees for his solar voltaic startup.
Back in March 2009, I hosted a radio hour with Dave The Brain Wave on blog talk radio that dealt with Solyndra and their novel technology of placing photo voltaic within a glass cylindrical tube instead of a flat panel. Dave and I basically said 30 months ago that these guys stood little chance against the competing technology of polysilicon based solar cells. We discussed learning rates and that companies that have innovative yet unique technology cannot rely on the communal learning of a broadly adopted technology and that Solyndra like the Sony Betamax will likely fail. Of course President Obama and Secretary Chu thought otherwise and will now face the questioning of the GOP. President Obama will address Congress next week on his plan for job creation. After his green jobs fiasco I wonder what the next debacle he is planning? Perhaps we should not put colors to jobs. Perhaps we should just talk about productive useful jobs. Perhaps the President should quickly get Secretary Chu to explain the second law of thermo to him. If Chu is not available perhaps the President should call the Green Machine who will explain the newly proposed fourth law of thermo. That is that "As governments start to spend vast amounts of money on ideas that violate the second law, then the entropy of the system approaches a maximum and Shift will Happen". Note the fourth law should not be mixed up with the third law that states "as temperature approaches absolute zero, entropy will approach a constant minimum".
The President is a lawyer and never did study thermo. He did get a Nobel Peace Prize for doing nothing a few weeks after entering office. His Secretary of Energy did win a Nobel Physics Prize for low temperature thermodynamics and Chu should have known better than to bet these billions on junk science. I have been asked why Chu sold out? Perhaps he was told to sell hope not thermo to the masses by his boss. Perhaps he simply loved the power of being Secretary of Energy of these Ignited States or perhaps he simply wanted the Chinese to win the economic battle and was happy to support junk science with billions of dollars while the Chinese erected polysilicon factories and beat the crap out of the likes of Solyndra. I cannot opine on whether Chu is an American Patriot but I can say for a guy who won a real Nobel Prize in low temperature physics he has been a joke of a Secretary of Energy and is right up there with Sam The Sham Bodman that hyped hydrogen and other piles of junk science. Both Chu and Bodman have science or engineering based PhDs. Both Chu and Bodman are flaming fools for what they have done to us. Chu certainly knows the third law of thermo as that was the basis for Nobel Prize in Physics. Now that the fourth law has been proclaimed, Chu should also be told it is time for "Shift to Happen". As for President Obama he should look in the mirror and see Jimmy Carter in the reflection.


Last week, Chris and I spent most of the week in Alabama. We went there to help get a new operation started with some of our trucks that were delivered on Wednesday morning. We also visited some of our current customers while we were there.


Procurement Minister Carla Qualtrough handed off the ceremonial keys to West Block and Parliament Hill's new Visitor Welcome Centre to House Speaker Geoff Regan at a Nov. 8 ceremony, followed by a media tour. Here's a look at the $863-million in renos West Block has undergone since it was closed in 2011, and the new $130-million underground visitors' centre. With Centre Block closing this winter for at least a decade of repairs, West Block is set to accommodate an interim House of Commons chamber, offices for the Speaker, House officers, and prime minister, and more.


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Cmentarz wojenny nr 302 – Żegocina – cmentarz z I wojny światowej zaprojektowany przez Franza Starka, znajdujący się we wsi Żegocina w województwie małopolskim, w powiecie bocheńskim, w gminie Żegocina. Jeden z ponad 400 zachodniogalicyjskich cmentarzy wojennych zbudowanych przez Oddział Grobów Wojennych C. i K. Komendantury Wojskowej w Krakowie.

Cmentarz znajduje się na cmentarzu parafialnym w jego centralnej części, w odległości około 400 m od kościoła. Zachowany i utrzymany jest w bardzo dobrym stanie. Zajmuje powierzchnię około 1950 m². W centralnej części cmentarza znajduje się pomnik stylizowany na ołtarz zwieńczony krzyżem z daszkiem. Na pomniku znajduje się tablica z napisem w języku niemieckim:

Znajduje się na nim 109 grobów pojedynczych oraz 54 zbiorowych. Pochowano w nich 276 żołnierzy poległych w 1914 roku:
 102 Rosjan m.in. z 50 Białostocki Pułk Piechoty, 292 Małoarchangielski Pułk Piechoty, 296 Griazowiecki Pułk Piechoty oraz z 147 Pułku Piechoty (147-й пехотный Самарский полк),
 22 Austriaków z 58 IR 83 IR, 295 IR, 18 LIR, 33 LIR, 35 LIR, K.U.K. F.J.B. 24, K.U.K. F.J.B. 29, 11 HIR,
 152 Niemców m.in. z 217, 218, 219, 220 i Pruskiego Rezerwowego Pułku Piechoty (Reserve Infanterie-Regiment (Preußische) Nr 217, 218, 219 i 220)

Bibliografia

Linki zewnętrzne
 Cmentarze z I Wojny Światowej. – Żegocina.
 Polegli wykaz Żegocina I (nr 302).
 Cmentarz wojskowy z I wojny światowej – nr 302.

Zachodniogalicyjskie cmentarze z I wojny światowej
Cmentarze z I wojny światowej w województwie małopolskim
Zabytki w powiecie bocheńskim
302
Żegocina

redpajama_set_name: RedPajamaWikipedia


Text by George Capsis. Photo by Maggie Berkvist.
This decades-old fence bulging into the path of Charles Street pedestrians is a relic of famed real estate scavenger William Gottlieb's "never fix anything" dictum and will soon disappear into seven-story condo cash.


We host the Bundu Expo Gauteng for the past seven years at the end of July 29 -31 2011.
Focus on the outdoors, hunting, fishing and camping and it is a great weekend getaway for the whole family.
The biggest man made dam in a double store Hall.
A special Ladies Lane has been arranged this year with body painting competition and Miss Bundu Competition.


text:
Play the Slav
By FM James Vigus
Everyman Chess
In the Preface, FM Vigus writes that he was inspired to refresh his Slav repertoire after the 2006 Kramnik - Topalov World Championship Match, in which the opening played a starring role.
This is a repertoire book and as such is selective with the chosen variations. The Chebanenko Variation (4...a6) is not covered.
The Introduction is very well written and sets out the author's stall very nicely. The basic repertoire is outlined and there's plenty of general Slav information, including a good look at pawn structures and other general considerations.
The main chapters are split thus:
The Sokolov Defence
The Dutch Variation: Introduction and 9 Qe2
The Dutch Variation: 9 Nh4 and Related Lines
Knight to the Right: 6 Nh4
The Tolush-Geller Gambit: 5 e4
Fifth Move Alternatives
The Errot
Quiet Queen Moves
The Argentinean Defence
The Exchange Variation
FM Vigus has branched out in new directions as regards nomenclature. The slightly off-beat way of meeting: 1 d4 d5 2 c4 c6 3 Nf3 Nf6 4 Nc3 dxc4 5 a4 Bf5 6 Ne5 Nbd7 7 Nxc4 with 7 …Nb6 (as opposed to the more common 7 …Qc7) is named The Sokolov Variation. 1 d4 d5 2 c4 c6 3 Nf3 Nf6 4 e3 Bg4 is named The Errot as it carries with similar themes as The Torre Attack, only in reverse. The Argentinean Defence refers to: 1 d4 d5 2 c4 c6 3 Nc3 dxc4
The bulwark of the repertoire is the old Dutch Variation:
1.d4 d5 2.c4 c6 3.Nf3 Nf6 4.Nc3 dxc4 5.a4 Bf5 6.e3 e6 7.Bxc4 Bb4
The sharpest try for White is the Tolush-Geller Gambit, which still needs careful attention, although theoretically it should struggle.
'It must not be underestimated and in many cases leads to very complex play. The computer age, however, is not kind to such speculative gambits and theoretically speaking White is on very precarious ground here.'
This is a critical position. FM Vigus recommends 13...Bd5! Here, as it: '..prepares …b5-b4 while enabling the queen's knight to come to c6, and shores up the e6-pawn'. The tactical nuances are all competently covered and should be analysed in conjunction this note: 'The most important advice is: don't panic!'
I particularly enjoyed the section on the Exchange Variation. It is clear that for The Slav to be a viable weapon for Black, rather than merely a holding device, then the second player must find some way to infuse life into position after 1 d4 d5 2 c4 c6 3 cxd5 cxd5. White usually enjoys a slight pull against symmetrical systems.
Here, considerable attention is devoted to a less obliging method of play.
1 d4 d5 2 c4 c6 3 cxd5 cxd5 4 Nc3 Nc6 5 Nf3 Nf6 6 Bf4 Nh5
'It is time to introduce the Russian Counterattack, a system for Black that has been unaccountably neglected in books on the Slav despite having served as the standard Soviet antidote to the Exchange for generations.'
With 6...Nh5 Black is hoping to impose his authority on the game rather than allow White to sit on a small edge. Indeed, as this book shows, the second player can even take advantage of the newly mobile f-pawn by heading for a Stonewall with f7-f5 before dropping the Knight back to f6.
The suggested repertoire is narrow but that's not such a bad thing; there is no doubting that the Slav is virtually fireproof and the lines given here are not at all likely to be blown away by any big novelties. Consequently, this opening book should enjoy a longer than average shelf-life.
The Greatest Ever Chess Opening Ideas
By IM Christoph Scheerer
The evolution of chess ideas is an interesting topic for a book and one that requires great research. The bibliography is very impressive, running to five full pages; the author has clearly left no stone unturned in his desire to unearth relevant stories.
All the standards are there, from the ancient tales of Ruy Lopez and Greco, Marshall's original Attack against Capablanca, the rise of the Hypermoderns, through to modern times and Kasparov's final over-the-board contributions in his retirement tournament.
Example number 1 shows the ancient Italian Game and over 430 years later, in chapter 50, 'Hot Off the Press', brings the history of the development of chess opening bang up to date with Topalov's bomb at Wijk aan Zee earlier this year:
Topalov - Kramnik
12 Nxf7!? with the final word still to be said.
Some stories are better known than others; all are highly readable and entertaining.
A classic example is the tale of the Gothenburg Variation of the Najdorf Sicilian - in which Najdorf, Panno and Pilnik took on the might of Spassky, Keres and Geller, all in the same round of the 1955 Interzonal tournament (with a full view of each other's games on the demonstration boards) is recounted in full.
1.e4 c5 2.Nf3 d6 3.d4 cxd4 4.Nxd4 Nf6 5.Nc3 a6 6.Bg5 e6 7.f4 Be7 8.Qf3 h6 9.Bh4 g5 10.fxg5 Nfd7
Geller led the way with 11 Nxe6! and the Soviets went on to record three victories.
Each chapter concludes with a rating, based on the following criteria:
Longevity Rating
Creativity Value
I'm not so sure that such a rating system works particularly well, or indeed that it is necessary. 5/5 is the most common rating although it is clearly indicated when the author believes an opening has seen better days. The Jaenisch Gambit is one such variation judged to be a thing of the past.
'We have reached a position in which it is difficult to find any compensation for the missing pawn.'
'I wonder if Radjabov will continue to employ the Jaenisch, and I'm more than curious to see what he has planned against the approach recommended in this chapter! I just think that the Jaenisch is not playable anymore…'
This book walks the line between presenting pure theory and a prose history of chess openings ('Theostorical' as the author has it). However, it keeps its balance very well and makes for a most interesting read, ideal for browsing, delightful for dipping into.
It's a big book - 368 pages - with plenty of prose to complement the chess analysis.
Pawn Sacrifice!
Winning at Chess the Adventurous Way
By IM Timothy Taylor
In the words of the author:
'…the subject of this book is pawn sacrifices that occur in the middlegame (and, rarely, in the ending). And as I look at the above sentence, I see that occur' is not the right word: these pawn sacrifices were played - decided on, in the heat of battle, with no textbook or computer assistance. Perhaps 'bravely ventured' is the best phrase! In every game in this book, at some point one of the players said to himself, 'I'm going to sacrifice this pawn - I don't see when or how I'm getting it back, but I like my position'. And then he boldly went for it!'
Genuine pawn sacrifices do seem rather rare in the games of amateurs. The same players who willingly sacrifice Minor pieces for an attack are often too afraid of losing endgames to give up a humble foot soldier.
Yet there are plenty of excellent examples of successful pawn sacifices in this book, featuring the finest of chess champions.
Pawn sacrifices tend to be associated mainly with opening gambits but in truth the typical scenarios are legion.
There are 12 main chapters, each covering a thematic type of pawn sacrifice, namely:
King Attack
Deflecting
Pawn Cracker
Obstructive
Two Bishops
Charge Up
The actual moment of the pawn sacrifice is discussed, with emphasis on the favourable omens detected by the player offering the booty. IM Taylor then checks the 'thoughts' of Fritz to see if the idea is approved by the mechanical mind before adding his own human assessment. Needless to say, strong disagreements between man and machine are rife. For example, in this famous position.
Tal - Tolush
Moscow 1957
After 16...bxc4 Tal played 17 Bb1! and the Bishop eventually bludgeoned its way to freedom along the b1-h7 diagonal. Fritz doesn't like the sacrifice at all, preferring to recapture on c4 either immediately or after an exchange on g7.
It may seem surprising to a lot of readers that the player making the most pawn sacrifices in this book is the Tigran Petrosian, who just edges out Mikhail Tal.
Petrosian - Najdorf
Bled 1961
White realises that only the Black Queen prevents a full-scale invasion, so he is prepared to sacrifice a pawn to ensure the exchange. 35 Qb6!! Qxb6 36 axb6 Rb8 37 Rc7 and even though the b-pawn fell, the floodgates had opened with decisive effect.
IM Taylor writes in a very enthusiastic manner. Exclamation marks abound, more so in the prose than in the game moves. This can become a trifle wearing at times.
The inclusion of a game by his wife in the final chapter could be seen as a further indulgence.
However, these are minor matters. The illustrative games and examples should open a few eyes towards the beauty of unexpected pawn sacrifices.
'The advent of chess machines has made many people leery of the speculative pawn sacrifice in general - however, in my opinion, that fact just makes such sacrifices even more dangerous! Your opponents probably have little practice in dealing with such boldness. I hope that this book helps you too (to) find your 'inner Tal'! '
Play 1 b4!
By IM Yury Lapshun and US Master Nick Conticello
Nick Conticello tells the story of how the book came into being in his five-page introduction and how the project stretched from an initial three week target to take three years.
'There is one outstanding reason to play the Sokolsky: it is virtually certain that your opponent will be unfamiliar with the positions that arise.'
This is a good point and equally an equally valid justification for playing any of the more unusual openings. I know from my own games I have a earned a paltry half point from my three encounters with 1 b4 - and two of my opponents were rated considerably lower than I was at the time.
1 b4 won't appeal to all players, as the authors readily admit:
'If you want to guarantee yourself a small advantage with no risk, the Sokolsky is not for you!'
Games by one of the lesser-quoted players form part of the basis of the book.
'Part of our mission in writing this book was to present the best of Sokolsky's work to the English-speaking chess world.'
'However, our primary purpose was to out forth Yury Lapshun's games and ideas, as he is` certainly one of the strongest players who regularly plays the Sokolsky.'
The main contents:
The Sokolsky Gambit
1 b4 e5 2 Bb2 Bxb4
Black plays …e5 and …d6
Queen's Indian Systems
Black plays …d5 and …e6
Black plays …d5 and …Bf5/Bg4
1...c6, 1...f5 and Unusual Moves
Of the 84 complete games, the majority feature Sokolsky, Katalymov and Lapshun but World Champions Alekhine, Spassky and Fischer can all be seen pushing the b4 pawn too (albeit in simultaneous games in the case of Fischer).
It is important for both players to keep an eye on transpositions. In several of the illustrative games the players end up in a standard English Opening and it could be possible to trick those commanding the Black pieces into a line of play they wouldn't normally adopt.
Strange positions abound. For instance, after 1 b4 d5 2 Bb2 Bg4 White plays 3 Qc1, unpinning the e-pawn.
Free spirits would enjoy the opportunity to play The Sokolsky Gambit:
1 b4 e5 2 Bb2 f6 3 e4 Bbx4 4 Bc4
…with f2-f4 coming next.
It's all interesting stuff and 1 b4 would doubtless make a good surprise weapon (possibly in the first round of Open tournaments, to make future opponents waste preparation time over the course of the weekend).
For further details of these and other Everyman products, please visit: http://www.everymanchess.com/
Missed a review? Pop along to my archive:http://marshtowers.blogspot.com/2007/12/chess-review-archive.html
Staunton Memorial
'Forcing Chess Moves': More Information

redpajama_set_name: RedPajamaCommonCrawl


If you like experimenting with unique taste of exotic game or enjoy trying new flavors you will want to try one of Alligator Bob's gift pack selections. Be assured that our Premium Meat snacks contain no fillers, extenders or animal fat additives, smoked in a real smoke house with natural hardwoods for that rich, smokehouse flavor. Always "Guaranteed Good" or your money back.
Due to International Trade Regulations we can only ship within the continental U.S.
Orders Placed in the AM (EST) Monday - Friday will be Shipped the same day!
FLORIDA RETAILERS MUST HAVE A FLORIDA TAX CERTIFICATE ON FILE WITH GATORBOBS OR YOU WILL BE CHARGED SALES TAX.
NOTE: If your order only contains one 30 piece box, you will be charged $54.90 and not receive the discounted price.
Are you looking for a great. unique, low cost, addition to our store or market? Then you will want to try one of Alligator Bob's pack selections. Choose from a variety of alligator snacks and our line of exotic game sticks. Be assured that our Premium Meat snacks contain no fillers, extenders or animal fat additives. Smoked in a real smoke house with natural hardwoods for that rich, smokehouse flavor. Always "Guaranteed Good" or your money back.
Buy Back Guarantee: Spoiled or expired merchandise returned within 120 days of purchase.


text:
Q: Boost::Process compile issues, had to modify lib to get it to work So I downloaded the latest Boost::Process library and attempted to compile and I end up with this:
libs/boost-1.55.0/build/include/boost/process/operations.hpp:85:36: error: 'filesystem_error' is not a member of 'boost::filesystem'
I did go ahead and check this file out, I removed that line (Along with the if statement ontop of it) and it compiles fine after the modification. The odd thing is that filesystem_error is in fact a member of boost::filesystem so I don't understand why it's complaining.
Is there any way to fix this without having to remove those lines? I really don't want to do a modification like that.
Here's the latest Boost::Process (I am using Boost 1.55.0 in case you wanted the version number).

A: You need to fix it like I did yesterday[1]:
In boost/process/operations.hpp change
#include <boost/filesystem/path.hpp>

into
#include <boost/filesystem/path.hpp>
#include <boost/filesystem/convenience.hpp>

Or just
#include <boost/filesystem.hpp>

The reason appears to be that path.hpp no includes the header for the error class indirectly

[1] okay, it was hidden in the 'amalgamate' on Coliru :)

redpajama_set_name: RedPajamaStackExchange


A bit late to the party – but new home lab!
I finally saved enough money to get myself a respectable homelab. Many people have had home labs for years, I'm a little late on that bandwagon. But better late than never. I spent a bit more than I wanted, but I cost-justified part of it by eliminating Dropbox. Dealing with our volume of home photos and videos is a continual frustrating timesink. We have been syncing our photos to Dropbox, but as I always preach to customers, sync is not backup. I have been manually storing zipfiles of monthly photos up in Glacier as emergency recovery, but I had fallen behind. Dropbox lost two months of our photos that I will never be able to get back. I decided that along with serving my homelab needs, any storage solution that I purchased absolutely had to support sync directly from my endpoints as well as to multiple clouds. Since I knew I was going to buy a NAS, this effectively eliminated VSAN as a contender for the homelab.
I spent a little time looking around, but decided I didn't want to deal with something like FreeNAS on a custom-built box. Many of my other fellow engineers are happy with their Synology devices, so that's what I went with.
2TB 5400RPM drives fit the bill nicely. I saved money and didn't put any storage inside the Shuttles, instead opting for boot-from-USB.
For my VM storage, I went with a 4-drive RAID-5 with a 256GB Flash read/write cache. I've ready blog entries both for and against the Synology cache. Some people says it makes no difference, some says it makes a huge difference. So far I'd say it makes a huge difference in VM performance.
The shuttle PCs only come with a single onboard NIC, to do anything useful I bought a quad port gigabit NIC for each host.
Here is my BOM. I bought everything new from Amazon except for the Cisco switch, which was about $100 on ebay. I was a little disappointed to find that Amazon's regular pricing was cheaper than any of our employee discounts. The total spend was much higher than I originally thought – but given the photo sync requirements, I think this was the best solution that satisfies my need to learn and my need to protect our irreplaceable photos.
Here it is all assembled. The kids even got in on the action, my oldest installed the RAM and CPU and my youngest installed the Quad NIC!


text:
Q: Find out the last updated record in my DB using MAX in CASE statement I have APPLICATIONSTATUSLOG_ID primary key field on my table.
In order to find out the last updated record in my DB and the MAX(APPLICATIONSTATUSLOG_ID) is presumed to be the most recent record.
I tried this code :
SELECT
    MAX(CASE WHEN MAX(d.ApplicationStatusLog_ID) = d.ApplicationStatusLog_ID THEN d.ApplicationStatusID END) AS StatusID,
FROM
    ApplicationStatusLog d

But I get error:

Msg 130, Level 15, State 1, Line 53 Cannot perform an aggregate function on an expression containing an aggregate or a subquery.

My table looks like
ApplicationID - ApplicationStatusID - ApplicationStatusLogID
10000           17                    100
10000           08                    101
10000           10                    102
10001           06                    103
10001           10                    104
10002           06                    105
10002           07                    106
My output should be:
10000 10
10001 10
10002 07
Please help me understand and resolve my problem.

A: If you want to just find the last updated row, given that it has max value in APPLICATIONSTATUSLOG_ID column. The query would be:
SELECT *
FROM ApplicationStatusLog
WHERE ApplicationStatusLog_ID = (SELECT MAX(ApplicationStatusLog_ID) FROM ApplicationStatusLog )

EDIT
So as you stated in comment, the query for it will be:
DECLARE @statusId INT
SELECT @statusId = STATUSID
FROM ApplicationStatusLog
WHERE ApplicationStatusLog_ID = (SELECT MAX(ApplicationStatusLog_ID) FROM ApplicationStatusLog )

EDIT 2:
The query as per your edit in question will be:
 WITH C AS
(
    SELECT ApplicationID,ApplicationStatusID,ApplicationStatusLogID, ROW_NUMBER() OVER (PARTITION BY ApplicationID ORDER BY ApplicationStatusLogID DESC) AS ranking
    FROM ApplicationStatusLog
)
SELECT ApplicationID,ApplicationStatusID
FROM C
WHERE ranking = 1

A: You can join same table twice like this:
select IT.JoiningID, JT.MAXAPPLICATIONSTATUSID FROM dbo.[Table] IT
    INNER JOIN (
        Select JoiningID, MAX (APPLICATIONSTATUSID) MAXAPPLICATIONSTATUSID
            FROM dbo.[Table]
            GROUP BY JoiningID
    ) JT ON IT.JoiningID = JT.JoiningID

Now you have MAXAPPLICATIONSTATUSID per ID so you can write what you wand based on MAXAPPLICATIONSTATUSID.

A: Without full query
SELECT
 x.StatusId
 ...
FROM  <Table> a
CROSS APPLY
(
 SELECT x.APPLICATIONSTATUSID as StatusId
 FROM  <Table> x
 HAVING MAX(APPLICATIONSTATUSLOG_ID) = a.APPLICATIONSTATUSLOG_ID
 GROUP BY x.APPLICATIONSTATUSID
)


In the Skeena, the ancient cycle continues, unbroken, for now. Steelhead and all five species of wild salmon bring abundant marine-derived nutrients to feed a rich ecosystem for wildlife populations including grizzly and black bear, wolverine, wolf, moose, deer, mountain goat and wild birds. They also sustain human communities, some of which have been here for thousands of years.
The Skeena is one of North America's last remaining intact watersheds, where there is more than $100 billion in industrial projects proposed. To avoid the fate of other systems which have been dammed, polluted, over-fished and paved over, we need comprehensive strategies to assess cumulative effects and encourage regional planning which protect our wild salmon ecosystems and the communities who depend on them.
Formed in 2007, SkeenaWild Conservation Trust is dedicated to making the Skeena River and nearby coastal communities a global model of sustainability where large human and salmon populations coexist. We are working with governments, First Nations, communities and individuals to sustain the long-term health and resilience of the wild salmon ecosystem.
Sustainability requires imagination, compassion and long-term balance. Salmon are the backbone of the diverse cultures, economy and ecosystem in the Skeena region. Protecting this iconic species is fundamental to maintaining and building a healthy watershed and local communities.
Our regionally-based organization is focused on the effective use of science, laws and convening to empower communities to protect and strengthen vulnerable salmon populations, improve management decisions and deepen people's connection with wild salmon.
Our approach is based on human and ecological sustainability, rigorous research and community input. It's simple – we support responsible development, whether it be mining, forestry, fisheries, or energy, that does not put salmon and communities at risk. This is done by using widely accepted scientific indicators to assess impacts.
Habitat and Species Protection: We provide support, leadership and education regarding development that threatens the health, resilience and diversity of wild salmon and the habitats upon which they depend. We facilitate dialogue among residents of the watershed and work with communities and all levels of government to increase understanding of impacts from existing and proposed industrial development, improve proposed projects, and provide solutions.
Science and Research: We conduct and sponsor quality science and research relating relevant ecological, economic and social issues. Catalyze and support planning processes which advocate for ecologic, economic and social sustainability and resiliency.
Sustainable Fisheries Management: We collaborate with First Nations and conservation partners to reduce impacts on small vulnerable stocks, develop and implement rebuilding plans.
Community Engagement: We strive to build relationships, create dialogue, solicit feedback, share stories and information. We do this through social and educational programs, annual events, and conversations in our communities. SkeenaWild supports the internationally recognized right of Indigenous people to free, prior and informed consent. We work hard to empower Skeena salmon communities to improve decision-making and long-term planning, and build a Skeena watershed regional identity. We believe a global model to sustain salmon can sustain all species.
SkeenaWild depends on support from a variety of revenue sources to carry out our science and research initiatives, habitat protection projects, community engagement and education programs.


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Refashioned Gifts & Accessories Archives - Wench.
Suits You Bottle Bags – made from suit jacket sleeves and ties – perfect for carrying wine bottles to parties – a must for the coming season!
Bottle Bags – Suits You!
Cynlluniwyd gan / Designed by Percolated Design © 2018 Wench.


On 31st October, the locomotive's oval buffers were removed and replaced with authentic 'original' style round buffers, which will match the new buffer beam 'skirts' to be fitted next.
This entry was posted in Project D6948 blog. Bookmark the permalink.


text:
With an API Platform, Reinvention Is Easy
Disruption is a core aspect of Uber's business model and identity. Like many other businesses in this era of digital transformation and the API platform, Uber rose to prominence by giving an old service model—the taxi—a tech-forward face-lift. Now that ride-sharing is the standard service model, disruption means growing within and beyond their space—such as through their recent partnership with Cargo, which provides Uber passengers snacks to purchase during a ride.
Uber's expansion into various side hustles is interesting on its face. But it also raises important questions for businesses wanting to make their own transformation. How can they benefit from partnering with third-party services? What problems come with total reinvention? And how can the right technology help?
What Can You Own?
Uber's story has a key takeaway for businesses. While the enterprise world's biggest players can conquer an amazing portfolio of goods—everything from home electronics and streaming video to shipping and logistics—Uber focuses more narrowly on evaluating the company's space and strengths to identify possibilities for strategic partnerships. These partnerships help them own their place in the market.
There is no shortage of real-world success stories like Uber's. Companies absorb others to upgrade some aspect of their capabilities without the legwork, expense, and toil of ground-up development. High-profile partnerships like the Uber-Cargo arrangement form when two companies wish to expand their presence or offerings without formally moving into the space of another. These businesses reap the benefits of growth without facing the challenges of full-on expansion.
That said, businesses don't have to agree to partnerships, acquisitions, or mergers to tap their strengths. Smartly selected technological tools—often provided by a third-party vendor partner—can promote equivalent growth while the first-party company maintains its independence. This is where API platforms come in; they're powerful, flexible tools that help businesses expand their technological capabilities with less effort and expense than the rigors of first-party development.
What Can the Right API Platform Do for You?
There's clearly a reason the term "disruption" has become a business mainstay. Anyone familiar with modern success traits understands that disruption has become necessary for advancement.
APIs have features that can help companies be disruptive:
Simplicity. Perhaps the defining top-level feature of an API platform is its ability to add flexible, powerful capability to a company's current or upcoming slate of technological products. For many companies, this is transformative enough on its own: With a communications-focused platform, for instance, the ability to quickly add features like in-app messaging or calling (instead of building the tools themselves, establishing carrier relationships across region, etc.) is the ability to revolutionize internal and customer-facing tools with speed.
Power. Even "minor" features on a customer's side require significant technical architecture on the back end. Hence, tools that add major capability with minimal effort provide a faster path to disruption and space ownership. In the example of communications APIs, this could mean leveraging the API provider's ability to carry messages over a network, letting customers or employees connect with others through the company's newly updated app.
Flexibility and scalability. Whether a business runs a small test of its expanded functionality or immediately rolls out to thousands of waiting users, APIs can help. They save the first party from the expenses and growing pains that come with start-fresh development.
Speaking of starting fresh, APIs offer another space-ownership perk for the companies using them. In almost all cases, they let companies avoid a total refresh or overhaul of their existing technical structure. At minimum, this reduces companies' risk (and expense and effort) when trying to expand, disrupt, or capture a larger part of the market based on recognized strengths and capabilities. Depending on the need and situation, a company could use APIs to avoid a complete redo of an app they're otherwise happy with. APIs could also save the company from expansive procedural changes on the human side and architectural expansion on the technical.
The Strategic Growth Potential of APIs
Ultimately, APIs can help businesses grow in any direction they wish (or that their circumstances allow). One company could leverage the technology to upgrade their internal communications capability, supporting the type of cross-location communication that helps any organization flourish. Another could use similar tools to expand in their space, bringing offerings similar to a competitor's without building a competing product from the ground up.
Equally important, though, is the technology's ability to support organizations as they look for realistic ways to disrupt. When treading water and rebuilding from scratch are both unattractive options, APIs can help.
Vonage staff


Our next Jeep Renegade nominee is Hlubi Mboya, and she gives us an exclusive titbit on her personal life. What possible task does Gareth give Hlubi to complete?
What are Tom Eaton's thought processes when he approaches topical events? Insightful and witty, just like he is on social media, and his columns.
We get hold of Fanie de Villiers, and get his take on the Allan Donald saga.


ETA 3 to 6 Weeks. The item "Seagate Cheetah 15K. 5 ST3146855LW 146GB 15K rpm 68pin Ultra 320 SCSI Hard Drive" is in sale since Monday, May 14, 2018. This item is in the category "Computer, Tablets & Netzwerk\Laufwerke & Speichermedien\Festplatten (HDD, SSD & NAS)\Interne Festplatten". The seller is "xiaojie1988416" and is located in HONG KONG.


We have Hong Kong to thank for the refrigerated minibar. A Hilton food and beverage director took on an experiment which helped the chain see a 5% increase in profits after adding a minibar to each of the 800 rooms in its Hong Kong property.
Perhaps it was the novelty of dispensing with room service, but the trend caught on. The high prices live on, but it seems that the profits were short-lived. Currently, minibar beverages account for just 0.2% of food and beverage revenue (double that to 0.4% if you add food revenue from minibars).
Hotels have struggled to find ways to make the minibar both relevant and a revenue-maker. Australian website traveler.com.au explains that minibars take up space, consume energy, and are a debt burden because of the time they rob from the staff to restock and monitor use.
Plus, guests say they just don't put much priority on the minibar anymore. MSN reports that InterContinental Hotels Group surveyed guests and discovered only 1% thought the minibar was the most important feature of a room, while 61% said it was WiFi. TripAdvisor survey respondents, meanwhile, placed the minibar dead-last in terms of importance.
Some hotels have taken an unrefrigerated approach to making the minibar palatable and profitable. You aren't tempted by what you can't see. Why not put those snacks – which don't need to be refrigerated – on a counter where they can be seen? Hotel News Now reports that companies are growing their offerings of technology incorporates digital tablets and displays to feature what's available inside the minibar.
Other hotels have opted to dispense completely with their minibars. They're replacing them with technology powered solutions like the Bartech eTray. It can be placed anywhere in a hotel guest room and uses infrared product detection. The website ehotelier.com reports that solutions like this can cost just a quarter of the price of a traditional refrigerated minibar.
Thanks to Millennials and their quest to extract experience from a hotel stay, the visitor industry has an opportunity to reposition the mission of the minibar. It can be the perfect place to share drinkable (and edible) examples of what's local.
GQ's earlier article points out that hotels looking to connect visitors with locality are stocking them with spirits created in the area. And, who cares if you've got the ingredients – do you know how to make a Manhattan? Hotels are connecting the dots, adding shakers and other cocktail accoutrements.
One of the most interesting minibar innovations was recently covered by the Los Angeles Times, which reports that the Standard International hotel chain plans to stock its minibars with CBD-infused gourmet gumdrops, created for them by the cannabis brand Lord Jones.
These new approaches show that there might be more for the minibar to offer when hotels seek ways to connect the role it plays in the guest experience. Can it provide convenience at a justifiable price? Perhaps, stocked with the right things. It might have a better opportunity helping a hotel communicate to a deeper understanding of the location and its history. Today's visitor is thirsty for more of it.
Learn more about how we use design to help shape experience.


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First Windows Phone 7.5 'Mango' phone, Fujitsu IS12T, released on KDDI au in Japan
By Chris Ziegler Aug 24, 2011, 11:28am EDT
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Microsoft has just dropped us a line to note that Fujitsu and KDDI au have released the IS12T in Japan, becoming the first Windows Phone 7.5 (codenamed "Mango") handset to hit retail anywhere in the world. It's a little surprising considering that the updated Marketplace for Mango was just opened to third-party developers yesterday -- buyers might have to wait a few days before 7.5-specific apps start becoming available -- and interestingly, Microsoft says "some of the experiences associated with Windows Phone 7.5 have yet to be fully enabled," though it doesn't lay out the full list of what's missing at this point. Hopefully, this means a final build suited for a more global launch is imminent, but the company isn't tipping its hand on that just yet.
Source: KDDI au, Fujitsu


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Grbić ist ein slawischer Familienname.

Namensträger

 Adrian Grbić (* 1996), kroatisch-österreichischer Fußballspieler
 Itana Grbić (* 1996), montenegrinische Handballspielerin
 Ivo Grbić (* 1996), kroatischer Fußballtorwart
 Nadja Grbić (* 1963), österreichische Linguistin
 Nikola Grbić (* 1973), serbischer Volleyballspieler
 Petar Grbić (* 1988), montenegrinischer Fußballspieler
 Vladimir Grbić (* 1970), serbischer Volleyballspieler

Familienname
Slawischer Personenname