Neuroprotection of Triptolide against Amyloid-Beta1-42-induced toxicity via the Akt/mTOR/p70S6K-mediated Autophagy Pathway.

Triptolide is a natural active compound that has significant neuroprotective properties and shows promising effects in the treatment of Alzheimer's disease (AD). Recent studies have shown that autophagy occurs in AD. In this study, we determined whether autophagy regulated by triptolide ameliorates neuronal death caused by amyloid-Beta1-42 (Aß1-42). We examined the effects of triptolide on cell viability, autophagy, apoptosis, and the protein kinase B/mammalian target of the rapamysin/70 kDa ribosomal protein S6 kinase (Akt/mTOR/p70S6K) signaling pathway in PC12 cells. The results indicated that triptolide treatment exhibited a cytoprotective effect against cell injury induced by Aß1-42. Triptolide also reduced apoptosis and enhanced cell survival by decreasing autophagosome accumulation and inducing autophagic degradation. Furthermore, our results also showed that activating the Akt/mTOR/p70S6K mechanism was one reason for the protection of triptolide. Triptolide treatment protected against Aß1-42-induced cytotoxicity by decreasing autophagosome accumulation, and inducing autophagic degradation in PC12 cells. These findings also suggest that the reduction of autophagosome accumulation observed in triptolide-treated cells was Akt/mTOR/p70S6K pathway dependent. Overall, triptolide exhibits a neuron protective effect and this study provides new insight into AD prevention and treatment.

Protective Effect of Vitamin C on Triptolide-induced Acute Hepatotoxicity in Mice through mitigation of oxidative stress.

Triptolide, a purified diterpenoid from the herb Tripterygium wilfordii Hook.f., was widely used to treat many diseases. However, the hepatotoxicity of triptolide limited its clinical use. Research showed oxidative stress played an important role in triptolide-induced liver injury. To investigate the effect of vitamin C, which was one of the most effective antioxidants, on triptolide-induced hepatotoxicity and its potential mechanism in mice. In the present study, acute liver injury was induced by intraperitoneal injection of triptolide and vitamin C was orally administered. The results showed treatment with vitamin C prevented the triptolide-induced liver injury by reducing the levels of aspartate transaminase from 286.86 to 192.48 U/mL and alanine aminotransferase from 746.75 to 203.36 U/mL. Histopathological changes of liver corresponded to the same trend. Furthermore, vitamin C also protected the liver against triptolide-induced oxidative stress by inhibiting the generation of malondialdehyde (2.22 to 1.49 nmol/mgprot) and hydrogen peroxide (14.74 to 7.19 mmol/gprot) and restoring the level of total superoxide dismutase (24.32 to 42.55 U/mgprot) and glutathione (7.69 to 13.03 µg/mgprot). These results indicated that vitamin C could protect against triptolide-induced liver injury via reducing oxidative stress, and vitamin C may pose a significant health protection in the clinical use of triptolide.

MKL1 inhibits cell cycle progression through p21 in podocytes.

BACKGROUND: The glomerular podocyte is a highly specialized cell type with the ability to ultrafilter blood and support glomerular capillary pressure. However, little is known about the genetic programs leading to this functionality or the final phenotype. RESULTS: In the current study, we found that the expression of a myocardin/MKL family member, MKL1, was significantly upregulated during cell cycle arrest induced by a temperature switch in murine podocyte clone 5 (MPC5) cells. Further investigation demonstrated that overexpression of MKL1 led to inhibition of cell proliferation by decreasing the number of cells in S phase of the cell cycle. In contrast, MKL1 knockdown by RNA interference had the opposite effect, highlighting a potential role of MKL1 in blocking G1/S transition of the cell cycle in MPC5 cells. Additionally, using an RT(2) Profiler PCR Array, p21 was identified as a direct target of MKL1. We further revealed that MKL1 activated p21 transcription by recruitment to the CArG element in its promoter, thus resulting in cell cycle arrest. In addition, the expression of MKL1 is positively correlated with that of p21 in podocytes in postnatal mouse kidney and significantly upregulated during the morphological switch of podocytes from proliferation to differentiation. CONCLUSIONS: Our observations demonstrate that MKL1 has physiological roles in the maturation and development of podocytes, and thus its misregulation might lead to glomerular and renal dysfunction.

Glycyrrhiza polysaccharides may have an antitumor effect in γδT cells through gut microbiota and TLRs/NF-κB pathway in mice.

Tumor immunotherapy can be a suitable cancer treatment option in certain instances. Here we investigated the potential immunomodulatory effect of oral glycyrrhiza polysaccharides (GCP) on the antitumor function of γδT cells in intestinal epithelial cells in mice. We found that GCP can inhibit tumor growth and was involved in the regulation of systemic immunosuppression. GCP administration also promoted the differentiation of gut epithelia γδT cells into IFN-γ-producing subtype through regulation of local cytokines in gut mucosa. GCP administration increased local cytokine levels through gut microbiota and the gut mucosa Toll-like receptors / nuclear factor kappa-B pathway. Taken together, our results suggest that GCP might be a suitable candidate for tumor immunotherapy, although further clinical research, including clinical trials, are required to validate these results.

The role of glycogen synthase kinase-3ß in glioma cell apoptosis induced by remifentanil.

The aim of malignant glioma treatment is to inhibit tumor cell proliferation and induce tumor cell apoptosis. Remifentanil is a clinical anesthetic drug that can activate the N-methyl-D-aspartate (NMDA) receptor. NMDA receptor signaling activates glycogen synthase kinase-3ß (GSK-3ß). Discovered some 32 years ago, GSK-3ß was only recently considered as a therapeutic target in cancer treatment. The purpose of this study was to assess whether remifentanil can induce the apoptosis of C6 cells through GSK-3ß activation. 3-(4,5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) was used to detect cell viability. Hoechst 33342 staining and flow cytometry were used to detect cell apoptosis. The effect of GSK-3ß activation was detected using a GSK-3ß activation assay kit and 4-benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione (TDZD-8), a potent and selective small molecule inhibitor of GSK-3ß. The MTT assay indicated that remifentanil induced C6 cell death in a concentration- and time-dependent manner. Hoechst 33342 staining and flow cytometry showed that remifentanil significantly induced C6 cell apoptosis. The measurement of GSK-3ß activation showed that remifentanil increased the cellular level of GSK-3ß. All of these toxic effects can be attenuated by treatment with TDZD-8. These results suggest that remifentanil is able to induce C6 cell apoptosis through GSK-3ß activation, which provides a basis for its potential use in the treatment of malignant gliomas.

Nanosized copper oxide induces apoptosis through oxidative stress in podocytes.

Nanosized copper oxide (nano-CuO) has been widely used in many fields. Recent studies have shown that nano-CuO has toxic effects on various organs, but the effects of nano-CuO on kidney remain unclear. The aim of this study was to assess whether nano-CuO can induce the apoptosis of podocytes. The result of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay indicated that nano-CuO decreased podocyte viability in a concentration-dependent and time-dependent manner. The apoptotic assay by flow cytometry showed that nano-CuO induced podocyte apoptosis significantly. Meanwhile, the result of flow cytometric assay gave a clear indication that nano-CuO increased reactive oxygen species (ROS) level. The measurement of superoxide dismutase (SOD) and malondialdehyde (MDA) showed that nano-CuO decreased SOD and increased MDA levels in podocytes. These results may provide basic information for the safe application of nano-CuO in the future.

Inflammatory signaling pathways in the treatment of Alzheimer's disease with inhibitors, natural products and metabolites (Review).

The intricate nature of Alzheimer's disease (AD) pathogenesis poses a persistent obstacle to drug development. In recent times, neuroinflammation has emerged as a crucial pathogenic mechanism of AD, and the targeting of inflammation has become a viable approach for the prevention and management of AD. The present study conducted a comprehensive review of the literature between October 2012 and October 2022, identifying a total of 96 references, encompassing 91 distinct pharmaceuticals that have been investigated for their potential impact on AD by inhibiting neuroinflammation. Research has shown that pharmaceuticals have the potential to ameliorate AD by reducing neuroinflammation mainly through regulating inflammatory signaling pathways such as NF­κB, MAPK, NLRP3, PPARs, STAT3, CREB, PI3K/Akt, Nrf2 and their respective signaling pathways. Among them, tanshinone IIA has been extensively studied for its anti­inflammatory effects, which have shown significant pharmacological properties and can be applied clinically. Thus, it may hold promise as an effective drug for the treatment of AD. The present review elucidated the inflammatory signaling pathways of pharmaceuticals that have been investigated for their therapeutic efficacy in AD and elucidates their underlying mechanisms. This underscores the auspicious potential of pharmaceuticals in ameliorating AD by impeding neuroinflammation.

[Hydrogen sulfide stimulates the development of rat glioblastoma].

OBJECTIVE: To address the hypothesis that hydrogen sulfide (H(2)S) is a functionally significant stimulator in the development of glioblastoma (GBM) and explore the mechanism of stimulation. METHODS: Forty adult Sprague-Dawley (SD) rats were given intracerebral injection of rat C6 glioma cell suspension, and an intraperitoneal injection of sodium hydrosulfide (NaHS), an exogenous H(2)S donor. The 40 rats were randomly divided into 4 groups of 10 rats in each: the control group, NaHS group, C6 glioma group (intracerebral implantation of C6 glioma cells) and C6-NaHS group (intracerebral implantation of C6 glioma cells and intraperitoneal injection of NaHS). Food and water were freely available during all phases of the experiment. Physical symptoms were observed and the tumor size was measured. Histological changes were examined by pathology. Immunohistochemical staining was used to analyze the expression of HIF-1α and integrated optical density (IOD) was used to determine the tumor microvessel density (MVD). The H(2)S content in the brain was measured. RESULTS: The physical symptoms of tumor-bearing rats became more serious after NaHS injection. The H(2)S level in the C6 glioma group was higher than that in the control group [(35.25 ± 1.03) nmol/g vs. (29.12 ± 0.94) nmol/g, P < 0.05], and the highest H(2)S level was found in the C6-NaHS group. The pathological examination showed that the implanted tumors were predominantly spheroid with a distinct border and no capsule could be detected. Neovascular proliferation was also observed. Foci of tumor necrosis, intratumoral hemorrhage, pseudopalisades and tumor cavity were clearly observed. The glioma cells had scant eosinophilic cytoplasm and enlarged hyperchromatic nuclei. All these phenomena were more markedly in the C6-NaHS group compared with that in other three groups. The mean tumor volume was significantly different between the C6 and C6-NaHS rats [(32.0 ± 6.9) mm(3) vs. (67.8 ± 11.9) mm(3), P < 0.001]. Immunohistochemical analysis exhibited that the hypoxia-inducible factor-1alpha (HIF-1α) and CD34 expression were significantly increased after the intraperitoneal injection of NaHS in the C6-NaHS rats (comparing the IOD between C6-NaHS group and C6 group, HIF-1α: 133 962.9 ± 451.4 vs. 38 569.8 ± 408.6, P < 0.001; CD34: 73 368.6 ± 404.8 vs. 14 570.6 ± 748.7, P < 0.001). Moreover, compared with the C6 group, there were higher MVD in the C6-NaHS group [(41.2 ± 7.9)/mm(2) vs. (97.0 ± 10.8)/mm(2), P < 0.001]. CONCLUSIONS: H(2)S serves as a stimulator in the development of rat glioblastoma and exogenous H(2)S strongly promotes the tumor growth. The stimulating mechanisms include the increase of HIF-1α expression and neovascular formation. H(2)S may be a significant regulator in the development of tumor.

Nanosized copper particles induced mesangial cell toxicity via the autophagy pathway.

Nanosized copper particles (nano Cu) have been incorporated into products in multiple industries, although studies have demonstrated that these particles are nephrotoxic. We investigated the cytotoxicity of nanosized copper particles on rat mesangial cells and measured rates of apoptosis, the expression of caspase-3, and generation of reactive oxygen species. We also measured autophagy through the acridine orange (AO) staining and expression of Beclin-1, microtubule-associated protein 1 light chain 3, and p62 to screen the underlying mechanism of toxicity. Nanosized copper particles inhibited mesangial cell viability, up-regulated the activity of caspase-3, and increased the rates of apoptosis and the generation of reactive oxygen species in a concentration-dependent manner. Exposure to nano Cu increased the formation of acidic vesicular organelles and the expression of Beclin-1, microtubule-associated protein 1 light chain 3, and p62, and treatment with an autophagy inhibitor reduced nephrotoxicity. This indicated that the autophagy pathway is involved in the toxicity induced by nanosized copper particles to mesangial cells. This finding can contribute to the development of safety guidelines for the evaluation of nanomaterials in the future.

Long non­coding RNA H19 inhibition ameliorates oxygen­glucose deprivation­induced cell apoptosis and inflammatory cytokine expression by regulating the microRNA­29b/SIRT1/PGC­1α axis.

As one of the earliest discovered long non­coding (lnc)RNAs, lncRNA H19 imprinted maternally expressed transcript (H19) participates in regulating ischemic stroke. The present study aimed to investigate the combined roles of lncRNA H19, microRNA (miR)­29b, silent mating­type information regulation 2 homolog 1 (SIRT1) and peroxisome proliferator­activated receptor­g co­activator­1α (PGC­1α) following ischemic stroke. lncRNA H19 expression levels in the middle cerebral artery occlusion (MCAO) mouse model and HT22 cells subjected to oxygen­glucose deprivation (OGD) were detected via reverse transcription­quantitative PCR (RT­qPCR). H19 small interfering RNA was used to knockdown H19 expression. Following OGD treatment, MTT, flow cytometry, ELISA, RT­qPCR and western blotting assays were performed to assess cell proliferation, cell apoptosis, inflammatory cytokine concentrations, and lncRNA H19, miR­29b, SIRT1, PGC­1α expression levels, respectively. In the present study, MCAO model mice and OGD­treated cells displayed significantly increased lncRNA H19 expression levels compared with sham mice and control cells, respectively. lncRNA H19 knockdown ameliorated OGD­induced cell apoptosis and increases in inflammatory cytokine concentrations. Furthermore, lncRNA H19 knockdown also attenuated OGD­mediated downregulation of miR­29b, SIRT1 and PGC­1α expression levels. Collectively, the results of the present study demonstrated that lncRNA H19 knockdown ameliorated OGD­induced cell apoptosis and increases in inflammatory cytokine concentrations by regulating miR­29b, SIRT1 and PGC­1α expression levels, which suggested the potential role of lncRNA H19 in ischemic stroke.

Effects of hydroxyapatite extract on rats with transient ischemia: Long-term potentiation and axon regeneration.

Hydroxyapatite (HA) has been extensively used as a reconstructive and prosthetic material for osseous tissue. The present study aimed to determine whether HA extract exerted effects on central nervous system injury following transient cerebral ischemia/reperfusion in rats. Male Wistar rats were treated with HA following bilateral common carotid artery clamping (two-vessel occlusion). The results demonstrated that treatment with HA extract attenuated the inhibition of long-term potential in a rat model of transient cerebral ischemia/reperfusion. Furthermore, HA extract improved axon regeneration, which was confirmed via the immunohistochemical analysis of growth associated protein 43 and glial fibrillary acidic protein. Taken together, the results of the present study provided preliminary evidence of the protective effect of HA on neuronal damage.

Overexpression of MicroRNA-16 Alleviates Atherosclerosis by Inhibition of Inflammatory Pathways.

BACKGROUND: Our previous study demonstrated that the expression of miR-16 was downregulated in the cell and animal models of atherosclerosis (AS), a main contributor to coronary artery disease (CAD). Overexpression of miR-16 inhibited the formation of foam cells by exerting anti-inflammatory roles. These findings indicated miR-16 may be an anti-atherogenic and CAD miRNA. The goal of this study was to further validate the expression of miR-16 in CAD patients and explore its therapeutic roles in an AS animal model. METHODS: A total of 40 CAD patients and 40 non-CAD people were prospectively registered in our study. The AS model was established in ApoE-/- mice fed a high-fat diet. The model mice were randomly treated with miR-16 agomiR (n = 10) or miR-negative control (n = 10). Hematoxylin-eosin staining was conducted for histopathological examination in thoracic aorta samples. ELISA and immunohistochemistry were performed to determine the expression levels of inflammatory factors (IL-6, TNF-α, MCP-1, IL-1ß, IL-10, and TGF-ß). qRT-PCR and western blotting were carried out to detect the mRNA and protein expression levels of PDCD4, miR-16, and mitogen-activated protein kinase pathway-related genes. RESULTS: Compared with the normal control, miR-16 was downregulated in the plasma and peripheral blood mononuclear cell of CAD patients, and its expression level was negatively associated with IL-6 and the severity of CAD evaluated by the Gensini score, but positively related with IL-10. Injection of miR-16 agomiR in ApoE-/- mice reduced the formation of atherosclerotic plaque and suppressed the accumulation of proinflammatory factors (IL-6, TNF-α, MCP-1, and IL-1ß) in the plasma and tissues but promoted the secretion of anti-inflammatory factors (IL-10 and TGF-ß). Mechanism analysis showed overexpression of miR-16 might downregulate target mRNA PDCD4 and then activate p38 and ERK1/2, but inactivate the JNK pathway. CONCLUSIONS: Our findings suggest miR-16 may be a potential diagnostic biomarker and therapeutic target for atherosclerotic CAD.

AMPK-dependent phosphorylation of HDAC8 triggers PGM1 expression to promote lung cancer cell survival under glucose starvation.

Cancer cells undergo metabolic reprogramming to sustain their own survival under an environment of increased energy demand; however, the mechanism by which cancer cells ensure survival under glucose deprivation stressed conditions remains elusive. Here, we show that deprivation of glucose, dramatically activated the glycogen pathway, accompanied by elevated phosphoglucomutase 1 (PGM1) expression. We further identified that AMP-activated protein kinase (AMPK) stimulated PGM1 expression by inducing histone deacetylase 8 (HDAC8) phosphorylation. Moreover, we demonstrated that glucose deprivation-induced AMPK activation stimulated the translocation of HDAC8 from the nucleus to the cytoplasm, consequently disrupting the binding between HDAC8 and histone 3. PGM1 expression was also found to be critical for lung cancer glycolysis, the oxidative pentose phosphate pathway, and oxidative phosphorylation under glucose deprivation conditions, and further led to the aberrant expression of metabolic enzymes involved in glucose metabolism mediated by ERK1/2. Finally, PGM1 was found to be highly expressed in lung cancer tissues from patients, which correlated with a poor prognosis. Taken together, these results revealed that AMPK activation by glucose deprivation leads to enhanced PGM1 expression, an essential component of the metabolic switch, to facilitate cancer progression, suggesting PGM1 as promising anti-cancer treatment targets.

Nanosized copper particles induced mesangial cell toxicity via the autophagy pathway

Nanosized copper particles (nano Cu) have been incorporated into products in multiple industries, although studies have demonstrated that these particles are nephrotoxic. We investigated the cytotoxicity of nanosized copper particles on rat mesangial cells and measured rates of apoptosis, the expression of caspase-3, and generation of reactive oxygen species. We also measured autophagy through the acridine orange (AO) staining and expression of Beclin-1, microtubule-associated protein 1 light chain 3, and p62 to screen the underlying mechanism of toxicity. Nanosized copper particles inhibited mesangial cell viability, up-regulated the activity of caspase-3, and increased the rates of apoptosis and the generation of reactive oxygen species in a concentration-dependent manner. Exposure to nano Cu increased the formation of acidic vesicular organelles and the expression of Beclin-1, microtubule-associated protein 1 light chain 3, and p62, and treatment with an autophagy inhibitor reduced nephrotoxicity. This indicated that the autophagy pathway is involved in the toxicity induced by nanosized copper particles to mesangial cells. This finding can contribute to the development of safety guidelines for the evaluation of nanomaterials in the future.

Triptolide attenuated injury via inhibiting oxidative stress in Amyloid-Beta25-35-treated differentiated PC12 cells.

BACKGROUND: Recently, an abnormal deposition of Amyloid-Beta (Aß) was considered the primary cause of the pathogenesis of Alzheimer's disease (AD). And how to inhibit the cytotoxicity is considered an important target for the treatment of AD. Triptolide (TP), a purified diterpenoid from the herb Tripterygium wilfordii Hook.f. (TWHF), has potential neuroprotective effects pertinent to disease of the nervous system. However, whether triptolide and its specific mechanisms have protective functions in differentiated PC12 cells treated with Aß25-35 remain unclear. AIMS: The purpose is to investigate the protective functions of triptolide in Aß25-35-stimulated differentiated PC12 cells. MAIN METHODS: In the study, we use 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, lactate dehydrogenase (LDH) assay, flow cytometry assay, immunohistochemical staining and Western blot to observe the effects of triptolide on cytotoxicity induced by Aß25-35 and its mechanism of oxidative stress. KEY FINDINGS: The result of MTT and LDH assay indicates that triptolide protected PC12 cells against Aß25-35-induced cytotoxicity. The flow cytometry assay shows that triptolide attenuated Aß25-35-induced apoptosis in differentiated PC12 cells. Meanwhile, the results give a clear indication that triptolide could downregulate generation of reactive oxygen species (ROS), hydrogen peroxide (H2O2) and malondialdehyde (MDA) induced by Aß25-35. The apoptotic process triggered by triptolide involved the up-regulation of the activity of superoxide dismutase (SOD). SIGNIFICANCE: The results suggest that triptolide may serve as an important role in the inhibition of the cell apoptosis induced by Aß and the decreased oxidative stress is a key mechanism in the protective effect of triptolide in AD.

Different expressions of large-conductance Ca2+-activated K+ channels in the mouse renal cortex and hippocampus during postnatal development.

Large-conductance Ca(+)-activated K(+) (BKCa) channels are widely distributed in a variety of cells and play a pivotal and specific role in many pathophysiological conditions. However, the function of BK(Ca) channels in the kidney cortex and hippocampus during the postnatal development has not received attention. In this study, to elucidate the role of BK(Ca) channels during the development, it is essential to establish the location and quantitation of expression of BK(Ca). The expressions of BK(Ca) were detected in the kidney and hippocampus on postnatal days (P) 1, 3, 5, 7, 14, 21, 28, and 49 by immunohistochemical and Western blot analysis. Our results showed that expressions of BK(Ca) channels were found in tubules and corpuscles at all time points. The expression was also observed at all developmental stages of the renal corpuscles, such as comma-shaped body, S-shaped body, renal corpuscles of stage III, and renal corpuscles of stage IV. During the development, the expression of BK(Ca) channels was decreased and the most prominent change of BK(Ca) protein level appeared between P14 and P21. In contrast, BK(Ca) channels were expressed in all regions of the hippocampus at every time point with the level increasing during the early development (P1 to P14). The findings of the present study suggest that BKCa channels play an important role during the postnatal development in both the renal cortex and hippocampus.

Triptolide Inhibited Cytotoxicity of Differentiated PC12 Cells Induced by Amyloid-Beta25₋35 via the Autophagy Pathway.

Evidence shows that an abnormal deposition of amyloid beta-peptide25-35 (Aß25-35) was the primary cause of the pathogenesis of Alzheimer's disease (AD). And the elimination of Aß25-35 is considered an important target for the treatment of AD. Triptolide (TP), isolated from Tripterygium wilfordii Hook.f. (TWHF), has been shown to possess a broad spectrum of biological profiles, including neurotrophic and neuroprotective effects. In our study investigating the effect and potential mechanism of triptolide on cytotoxicity of differentiated rat pheochromocytoma cell line (the PC12 cell line is often used as a neuronal developmental model) induced by Amyloid-Beta25-35 (Aß25-35), we used 3-(4, 5-dimethylthiazol-2-yl)-2, 5- diphenyltetrazolium bromide (MTT) assay, flow cytometry, Western blot, and acridine orange staining to detect whether triptolide could inhibit Aß25-35-induced cell apoptosis. We focused on the potential role of the autophagy pathway in Aß25-35-treated differentiated PC12 cells. Our experiments show that cell viability is significantly decreased, and the apoptosis increased in Aß25-35-treated differentiated PC12 cells. Meanwhile, Aß25-35 treatment increased the expression of microtubule-associated protein light chain 3 II (LC3 II), which indicates an activation of autophagy. However, triptolide could protect differentiated PC12 cells against Aß25-35-induced cytotoxicity and attenuate Aß25-35-induced differentiated PC12 cell apoptosis. Triptolide could also suppress the level of autophagy. In order to assess the effect of autophagy on the protective effects of triptolide in differentiated PC12 cells treated with Aß25-35, we used 3-Methyladenine (3-MA, an autophagy inhibitor) and rapamycin (an autophagy activator). MTT assay showed that 3-MA elevated cell viability compared with the Aß25-35-treated group and rapamycin inhibits the protection of triptolide. These results suggest that triptolide will repair the neurological damage in AD caused by deposition of Aß25-35 via the autophagy pathway, all of which may provide an exciting view of the potential application of triptolide or TWHF as a future research for AD.

Developmental changes in the expression and function of TRPC6 channels related the F-actin organization during differentiation in podocytes.

The transient receptor potential canonical (TRPC) 6 channel is an important ion channel located in podocytes, which plays an essential role in regulating calcium homeostasis of the cell signaling. Podocytes are specialized, terminally differentiated cells surrounding glomerular capillaries, and are the subject of keen interest because of their key roles in kidney development and disease. Here we wonder whether TRPC6 channels undergo developmental changes in the expression and function during the podocyte differentiation, and whether they contribute to the maturation of podocytes. Using morphological, immunohistochemical and electrophysiological techniques, we investigated the development of distribution and expression of TRPC6 in conditionally immortalized mouse podocyte cell line. Our results showed that the distribution of TRPC6 channels changed with the maturity of podocyte differentiation. The fluorescent intensity of TRPC6 on cell surface increased, which was accompanied by a corresponding increase in the density of current flowing through the channels. TRPC6 inhibition by TRPC6 siRNA or SKF-96365, a blocker or TRP cation channels, resulted in F-actin cytoskeleton disruption only on the developmental stage of podocytes. These results strongly support the conclusion that TPRC6 is an essential component of the slit diaphragm and is required for development of glomerulus.

Developmental changes of BKCa channels depend on differentiation status in cultured podocytes.

The podocyte is a remarkable cell type, which encases the capillaries of the kidney glomerulus. Podocytes are of keen interests because of their key roles in kidney development and disease. Large-conductance Ca(2+)-activated K(+) channels (BKCa channels) are important ion channels located in podocytes and play the essential role in regulating calcium homeostasis cell signaling. In this research, we studied the undergoing developmental changes of BKCa channels and their contribution to functional maturation of podocytes. Our results showed that the distribution of BKCa channels changed with the maturity of differentiation in a conditionally immortalized mouse podocyte cell line. Additionally, the increase of BKCa channel protein expression was detected by immunofluorescence staining with confocal microscopy in podocytes, which was consistent with the increase in the current density of BKCa channels examined by whole-cell patch-clamp technique. Our results suggested that the developmental changes of BKCa channels may help podocytes adapt to changes in pressure gradients occurring in physiological conditions. Those findings may have implications for understanding the physiology and development of kidney and will also serve as a baseline for future studies designed to investigate developmental changes of ion channel expression in podocytes.

Protective effects of leukemia inhibitory factor against oxidative stress during high glucose-induced apoptosis in podocytes.

Leukemia inhibitory factor (LIF) is a pleiotropic glycoprotein belonging to the interleukin-6 family of cytokines. In kidney, LIF regulates nephrogenesis, involves in tubular regeneration, responds to pro- and anti-inflammatory stimuli, and so on. LIF also plays an essential role in protective mechanisms triggered by preconditioning-induced oxidative stress. Although LIF shows a wide range of biologic activities, effects of LIF on high glucose-induced oxidative stress in podocytes remain unclear. The aim of the study was to assess whether LIF can attenuate high glucose-induced apoptosis in podocytes. The result of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay indicated that LIF protected podocytes against high glucose-induced cytotoxicity. The flow cytometry assay showed that LIF attenuated high glucose-induced apoptosis in podocytes. Meanwhile, the result of flow cytometric assay gave the clear indication that LIF decreased high glucose-induced elevated level of reactive oxygen species (ROS). The measurement of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, superoxide dismutase (SOD), malondialdehyde (MDA), and caspase-3 activity levels showed that LIF attenuated the high glucose-induced decreased level of SOD and elevated level of NADPH oxidase, MDA and caspase-3 activity. These results may provide potential therapy for diabetic nephropathy in the future.