[Zinc deficiency induces exfoliation of round spermatids by decreasing the expressions of TGF-ß1 and FAK in the mouse testis].

OBJECTIVE: To investigate the mechanisms of zinc deficiency inducing spermatogenic disorders. METHODS: Forty 4-week-old CD-1 male mice were randomly divided into two groups of equal number: experimental and control, the former fed on a low-zinc diet and the latter on a normal diet, both for 5 weeks. Then all the mice were sacrificed and their testes and epididymides harvested for detection of the concentration of zinc ion in the testis by atomic absorption spectrophotometry, observation of the histopathological changes in the testis and epididymis by HE staining, examination of the properties of the exfoliated cells by dual immunofluorescence staining and determination of the expressions of ZO-1, FAK, TGF-ß1, TGF-ß2, TNF-α and Par6 proteins in the testicular tissue by Western blot. RESULTS: The concentration of zinc ion in the testis was significantly lower in the experimental than in the control group (ï¼»140.59 ± 16.22ï¼½ vs ï¼»218.44 ± 31.29ï¼½ µg/g, P < 0.05). HE staining showed normal testicular tissue structure, dense seminiferous tubules and intact seminiferous epithelium, with clear and orderly arrangement of spermatogenic cells at all levels in the control group. The ratio of the abnormal seminiferous tubules to the total number was 0.01 ± 0.01. The mice in the experimental group, however, exhibited degeneration of seminiferous epithelium, reduced number of spermatids, vacuolated cytoplasm of Sertoli cells, occluded seminiferous tubules, and a remarkably larger number of abnormal seminiferous tubules than that in the control (0.75 ± 0.04 vs 0.25 ± 0.04, P < 0.01). Exfoliated cells were observed in the abnormal tubules and the caput, corpus and cauda of the epididymis in the experimental group, which were shown to be immature round spermatids in H1T2 and TRA54 dual-immunofluore-scence staining. Western blot manifested that the protein expression of ZO-1 was 0.904 ± 0.052 vs 1.130 ± 0.054 in the experimental and control groups, that of Par6 was 0.129 ± 0.049 vs 0.145 ± 0.047, that of TGF-ß2 was 0.116 ± 0.047 vs 0.142 ± 0.048, and that of TNF-α was 0.469 ± 0.022 vs 0.458 ± 0.023, with significant decreases in the former group as compared with the latter in the levels of FAK (0.144 ± 0.047 vs 0.219 ± 0.048, P < 0.05) and TGF-ß1 (0.024 ± 0.058 vs 0.586 ± 0.048, P < 0.01). CONCLUSIONS: Zinc deficiency can induce histopathological changes in the testis of the mouse, leading to exfoliation of round spermatids, in which FAK and TGF-ß1 may play an essential contributive role.

Coccomyxa Gloeobotrydiformis Polysaccharide Inhibits Lipopolysaccharide-Induced Inflammation in RAW 264.7 Macrophages.

BACKGROUND/AIMS: Inflammation plays a vital role in the etiology and pathogenesis of chronic noncommunicable diseases (NCDs), which are the leading health issues throughout the world. Our previous studies verified the satisfactory therapeutic effects of Coccomyxa gloeobotrydiformis (CGD) polysaccharide on several NCDs. In this study, we aimed to investigate the anti-inflammatory effects of CGD polysaccharide, and the corresponding molecular mechanisms, on lipopolysaccharide (LPS)-induced inflammation in RAW264.7 cells. METHODS: A viability assay and a lactate dehydrogenase (LDH) assay were used to measure the cytotoxic effects of CGD polysaccharide on LPS-stimulated RAW264.7 cells. To investigate the potential anti-inflammatory mechanisms of CGD polysaccharide in LPS-stimulated RAW264.7 cells, nitric oxide (NO) production was determined using a NO assay and the expression of inflammatory mediators (PGE2, iNOS and COX-2), inflammatory cytokines (TNF-α, IL-6, IL-1ß and IL-10) and inflammation-related signaling pathways (the MAPK/NF-κB, PI3K/AKT/JNK, JAK/STAT and Nrf2/HO-1pathways) were observed by western blotting. The translocation of NF-κB p65 was also observed using an immunofluorescent assay. RESULTS: CGD polysaccharide significantly inhibited LPS-induced NO production and PGE2 expression by reducing the expression of iNOS and COX-2. It also suppressed the expression of the pro-inflammatory cytokines TNF-α, IL-6 and IL-1ß, and up-regulated the expression of the anti-inflammatory cytokine IL-10. Further experiments demonstrated that CGD polysaccharide could inhibit inflammatory signaling pathways (the MAPK/NF-κB, PI3K/AKT/JNK and JAK/STAT pathways). At the same time, it enhanced the anti-inflammatory pathway Nrf2/HO-1. In addition, CGD polysaccharide did not display any cytotoxic effects, even at a high concentration. CONCLUSION: Taken together, the results suggest that CGD polysaccharide significantly inhibits LPS-induced inflammation in RAW264.7 cells. This effect lies in its regulatory effects on the signaling pathways MAPK/ NF-κB, PI3K/AKT/JNK, JAK/STAT and Nrf2/HO-1.Our findings reveal that CGD polysaccharide has the potential to be used as a relatively safe and effective drug as part of the treatment of NCDs.

Neuroprotective Effect of ZnT3 Knockout on Subarachnoid Hemorrhage.

BACKGROUND: The pathophysiology of early brain injury (EBI) after subarachnoid hemorrhage (SAH) is poorly understood. The present study evaluates the influence of zinc transporter 3 (ZnT3) knockout and the depletion of vesicular zinc on EBI. METHODOLOGY: SAH was induced in ZnT3 KO mice by internal carotid artery perforation. The changes in behavior were recorded at 24 hours after SAH. Hematoxylin-eosin, Nissl and TUNEL staining were performed to evaluate neuronal apoptosis. Data from mice with a score of 8-12 in intracerebral bleeding (i.e. moderate SAH), were analyzed. RESULTS: The degree of SAH-induced neuronal injury was directly correlated to the amount of blood lost, which in turn was negatively reflected in their behavior. The Wild Type (WT)-SAH group behaved poorly when compared to the knockout (KO)-SAH mice and their poor neurological score was accompanied by an increase in the number of apoptotic neurons. Conversely, the improvement of behavior in the KO-SAH group was associated with a marked reduction in apoptotic neurons. CONCLUSIONS: These results suggest that ZnT3 knockout may have played a vital role in the attenuation of neuronal injury after SAH and that ZnT3 may prove to be a potential therapeutic target for neuroprotection in EBI.

Protective Effect of Znt7 on High Glucose-Induced Epithelial-to-Mesenchymal Transition in Renal Tubular Epithelial Cells.

BACKGROUND/AIMS: Evidence from our and other groups has demonstrated that zinc transporter 7 in SLC30 family (ZnT7) inhibited epithelial-to-mesenchymal transition (EMT) and apoptosis in rat peritoneal mesothelial cells (RPMCs) under high glucose (HG) concentration. In the present study, we investigated the effect of ZnT7 on EMT of renal tubular epithelial cells (RTECs) in an in vitro model of diabetic nephropathy (DN). METHODS: A dual-fluorescent staining protocol was used for detection of ZnT7 in a normal rat kidney tubular epithelial cell line (NRK-52E cells). EMT was induced with HG (30 mM). NRK-52E cells were transfected with plasmids codifying for hZnT7-EGFP and interfering RNA for determination of the effect of ZnT7 over-expression and silencing, respectively. Expression of ZnT7, activation of the MAPK/ERK and TGF-ß/Smad pathways were analyzed with by means of Western blot. RESULTS: ZnT7 was localized in the perinuclear region and Golgi apparatus. In HG-induced EMT of NRK-52E cells, ZnT7 was up-regulated. Over-expression of ZnT7 led to inhibition of HG-induced EMT, while knock-down of ZnT7 increased EMT. Furthermore, knock-down of ZnT7 and increased HG-induced EMT was accompanied by activation of the MAPK/ERK and TGF-ß/Smad pathways. CONCLUSION: The present study provides evidence that ZnT7 has a protective effect over EMT of RTECs in DN and suggests that the inhibition of HG-induced EMT may be achieved through the MAPK/ERK and TGF-ß/Smad pathways. Thereby, ZnT7 could be a potential target for translation medicine and prevention program in DN.

Sorafenib versus sunitinib as first-line treatment agents in Chinese patients with metastatic renal cell carcinoma: the largest multicenter retrospective analysis of survival and prognostic factors.

BACKGROUND: To compare the efficacy of sorafenib and sunitinib with regard to overall survival (OS) and progression free survival (PFS) in Chinese patients with metastatic renal cell carcinoma (mRCC). METHODS: A multicenter, retrospective study was performed to elucidate the relationship between clinical variables and prognosis comparing sorafenib and sunitinib as first-line treatment agents in Chinese patients with mRCC. Between September 2006 and December 2014, 845 patients received either sorafenib (400 mg bid; n = 483) or sunitinib (50 mg q.d; n = 362). The primary end point was OS and PFS. RESULTS: The percentage of patients with low and moderate risk according to Memorial Sloan-Kettering Cancer Centre (MSKCC) score was significantly higher in sunitinib group, and that with high risk was significantly higher in sorafenib group (15.1 vs. 5.2%; p < 0.001). Median OS was similar in sorafenib and sunitinib group (24 vs. 24 months; p = 0.298). Sorafenib group exhibited higher mPFS compared to sunitinib group (11.1 vs. 10.0 months; p = 0.028). Treatment (sorafenib vs sunitinib), pathology, Eastern Cooperative Oncology Group (ECOG) performance status, MSKCC scores, Heng's criteria of risk, and number of metastases were identified as significant predictors for OS and along with liver metastasis for PFS. Clinical outcomes in terms of mOS was significantly better with sorafenib in patients ≥65 years of age (p = .041), ECOG 0 (p = 0.0001), and median MSKCC risk score (p = 0.008). CONCLUSIONS: Sorafenib and sunitinib are both effective in treating mRCC. However, sorafenib might be more effective in elderly patients (≥65 years) and in patients with an ECOG status of 0, classified under MSKCC moderate risk.

Effect of zinc deficiency on mouse renal interstitial fibrosis in diabetic nephropathy.

There is emerging evidence that tubulointerstitial fibrosis is the final common pathway of the majority of chronic progressive renal diseases, including diabetic nephropathy (DN). Zinc, an essential dietary element, has been suggested to be important for a number of protein functions during fibrosis in vivo and in vitro. However, the effect of zinc deficiency (ZnD) on renal interstitial fibrosis in DN remains unclear. The present study investigated the effect and the underlying mechanisms of ZnD on renal interstitial fibrosis during DN using an streptozotocin­induced model of diabetes with immunofluorescence staining and western blot analysis. The present study identified that dietary zinc restriction significantly decreased zinc concentrations in the plasma and mouse kidney. ZnD enhanced albuminuria and extracellular matrix protein expression, associated with diabetic renal interstitial fibrosis by activation of renal interstitial fibroblasts and regulation of the expression of fibrosis­associated factors, which may be mediated by the activation of fibroblasts via the TGF­ß/Smad signaling pathway. The data indicates that ZnD serves an important role in the pathogenic mechanisms of renal interstitial fibrosis during the development of DN.

Protective Effects of Berberine on Renal Injury in Streptozotocin (STZ)-Induced Diabetic Mice.

Diabetic nephropathy (DN) is a serious diabetic complication with renal hypertrophy and expansion of extracellular matrices in renal fibrosis. Epithelial-to-mesenchymal transition (EMT) of renal tubular epithelial cells may be involved in the main mechanism. Berberine (BBR) has been shown to have antifibrotic effects in liver, kidney and lung. However, the mechanism of cytoprotective effects of BBR in DN is still unclear. In this study, we investigated the curative effects of BBR on tubulointerstitial fibrosis in streptozotocin (STZ)-induced diabetic mice and the high glucose (HG)-induced EMT in NRK 52E cells. We found that BBR treatment attenuated renal fibrosis by activating the nuclear factor-erythroid 2-related factor 2 (Nrf2) signaling pathway in the diabetic kidneys. Further revealed that BBR abrogated HG-induced EMT and oxidative stress in relation not only with the activation of Nrf2 and two Nrf2-targeted antioxidative genes (NQO-1 and HO-1), but also with the suppressing the activation of TGF-ß/Smad signaling pathway. Importantly, knockdown Nrf2 with siRNA not only abolished the BBR-induced expression of HO-1 and NQO-1 but also removed the inhibitory effect of BBR on HG-induced activation of TGF-ß/Smad signaling as well as the anti-fibrosis effects. The data from present study suggest that BBR can ameliorate tubulointerstitial fibrosis in DN by activating Nrf2 pathway and inhibiting TGF-ß/Smad/EMT signaling activity.

SNF5 is Involved in Suppression of Hepatocellular Carcinoma Progression via TGF-Beta 1 Signaling.

SNF5 (SMARCB1/INI1/BAF47), a core subunit of SWI/SNF complex, has been reported to modulate cell proliferation and apoptosis. Genetic evidence has suggested that SNF5 participates in tumor suppression. However, the detailed biological function and underlying mechanisms of SNF5 in hepatocellular carcinoma (HCC) progression remain unclear. Here, SNF5 expression reduction in HCC tissues compared with the adjacent non-cancerous tissues has been demonstrated. Importantly, the results showed that reduced SNF5 expression has a strong correlation with worse overall survival of HCC patients. The data demonstrated that knockdown of SNF5 significantly promoted cell growth and migration in Hep3B and HCCLM3 cell lines. Interestingly, it was found that SNF5 suppressed transforming growth factor-ß1 (TGF-ß1) expression, and SNF5 mRNA expression was negatively correlated with TGF-ß1 in HCC tissues. Furthermore, depletion of SNF5 attenuated the sensitivity of HCC cells to sorafenib. Thus, the data suggested that SNF5 may participate in HCC suppression, and reduced expression of SNF5 correlates with the poor differentiation and prognosis of HCC, indicating that SNF5 might be an important prognostic biomarker and promising therapeutic target for HCC. Anat Rec, 299:869-877, 2016. © 2016 Wiley Periodicals, Inc.

A potential role for zinc transporter 7 in testosterone synthesis in mouse Leydig tumor cells.

Previous studies have demonstrated that zinc (Zn) is an essential trace element which is involved in male reproduction. The zinc transporter (ZnT) family, SLC30a, is involved in the maintenance of Zn homeostasis and in mediating intracellular signaling events; however, relatively little is known regarding the effect of ZnTs on testosterone synthesis. Thus, in the present study, we aimed to determine the effect of Zn transporter 7 (ZnT7) on testosterone synthesis in male CD-1 mice and mouse Leydig cells. The findings of the present study revealed that the concentrations of Zn in the testes and Leydig cells were significantly lower in mice fed a Zn-deficient diet compared with the control mice fed a Zn-adequate diet. In addition, ZnT7 was principally expressed and colocalized with steroidogenic acute regulatory protein (StAR) in the Leydig cells of male CD-1 mice. ZnT7 expression was downregulated in the mice fed a Zn-deficient diet, which led to decreases in the expression of the enzymes involved in testosterone synthesis namely cholesterol side­chain cleavage enzyme (P450scc) and 3ß-hydroxysteroid dehydrogenase/D5-D4 isomerase (3ß-HSD) as well as decreased serum testosterone levels. These results suggested that Znt7 may be involved in testosterone synthesis in the mouse testes. To examine this hypothesis, we used the mouse Leydig tumor cell line (MLTC-1 cell line) in which the ZnT7 gene had been silenced, in order to gauge the impact of changes in ZnT7 expression on testosterone secretion and the enzymes involved in testosterone synthesis. The results demonstrated that ZnT7 gene silencing downregulated the expression of StAR, P450scc and 3ß-HSD as well as progesterone concentrations in the human chorionic gonadotrophin (hCG)-stimulated MLTC-1 cells. Taken together, these findings reveal that ZnT7 may play an important role in the regulation of testosterone synthesis by modulating steroidogenic enzymes, and may represent a therapeutic target in testosterone deficiency.

Berberine activates Nrf2 nuclear translocation and inhibits apoptosis induced by high glucose in renal tubular epithelial cells through a phosphatidylinositol 3-kinase/Akt-dependent mechanism.

Apoptosis of tubular epithelial cells is a major feature of diabetic kidney disease, and hyperglycemia triggers the generation of free radicals and oxidant stress in tubular cells. Berberine (BBR) is identified as a potential anti-diabetic herbal medicine due to its beneficial effects on insulin sensitivity, glucose metabolism and glycolysis. In this study, the underlying mechanisms involved in the protective effects of BBR on high glucose-induced apoptosis were explored using cultured renal tubular epithelial cells (NRK-52E cells) and human kidney proximal tubular cell line (HK-2 cells). We identified the pivotal role of phosphatidylinositol 3-kinase (PI3K)/Akt in BBR cellular defense mechanisms and revealed the novel effect of BBR on nuclear factor (erythroid-derived 2)-related factor-2 (Nrf2) and heme oxygenase (HO)-1 in NRK-52E and HK-2 cells. BBR attenuated reactive oxygen species production, antioxidant defense (GSH and SOD) and oxidant-sensitive proteins (Nrf2 and HO-1), which also were blocked by LY294002 (an inhibitor of PI3K) in HG-treated NRK-52E and HK-2 cells. Furthermore, BBR improved mitochondrial function by increasing mitochondrial membrane potential. BBR-induced anti-apoptotic function was demonstrated by decreasing apoptotic proteins (cytochrome c, Bax, caspase3 and caspase9). All these findings suggest that BBR exerts the anti-apoptosis effects through activation of PI3K/Akt signal pathways and leads to activation of Nrf2 and induction of Nrf2 target genes, and consequently protecting the renal tubular epithelial cells from HG-induced apoptosis.

Zinc Attenuates Tubulointerstitial Fibrosis in Diabetic Nephropathy Via Inhibition of HIF Through PI-3K Signaling.

Evidence has demonstrated that hypoxia may have a central pathogenic mechanism in the development of diabetic nephropathy (DN). Epithelial-to-mesenchymal transition (EMT) of mature tubular epithelial cells in kidney is a contributor to the renal accumulation of matrix protein in DN and is highly associated with the progression of tubulointerstitial fibrosis. Zinc (Zn) has anti-fibrosis effects in liver and lungs. In the present study, we aimed to investigate the effect of Zn on renal tubulointerstitial fibrosis especially under hypoxic conditions and its association with DN. We found that Zn treatment blockaded tubular EMT and attenuated renal tubulointerstitial fibrosis by downregulation of hypoxia-inducible factor alpha (HIF-1α) in the kidneys of diabetic streptozotocin-treated mice. High glucose (HG)/hypoxic conditions stimulated EMT in renal tubular cells as indicated by the significant decrease in epithelial marker E-cadherin and ZO-1 while the increase in mesenchymal markers α-smooth muscle actin (α-SMA). Zn supplement mainly prevented HG/hypoxic-induced HIF-1α accumulation and EMT marker changes. In co-treatment Zn with PI3K/Akt/GSK-3ß signaling pathway, inhibitor LY294002 prevented HG/hypoxic-induced HIF-1α increase and EMT changes, suggesting that Zn may mediate HG/hypoxic-induced EMT through PI3K/Akt/GSK-3ß pathway. Therefore, we concluded that Zn had an important anti-fibrosis role under HG/hypoxic conditions, and a novel mechanism contributing to Zn protection on renal tubular epithelial cells from HG/hypoxia-induced EMT through activation of PI3K/Akt/GSK-3ß signaling pathway, which subsequently leads to the downregulation of the expression of HIF-1α.

Effect of zinc on high glucose-induced epithelial-to-mesenchymal transition in renal tubular epithelial cells.

Zinc (Zn) as an essential dietary element has been indicated in a number of protein functions in the prevention of numerous types of epithelial-to-mesenchymal transition (EMT)-driven fibrosis in vivo. However, relatively little is known regarding its effect in the EMT of the renal tubular epithelial cells, which play an important role in renal tubulointerstitial fibrosis and is an important component of the renal injury that is associated with diabetic nephropathy. The present study investigated the effect of Zn on the high glucose (HG)-induced EMT in a normal rat kidney tubular epithelial cell line (NRK-52E cells) and the underlying molecular mechanisms by immunofluorescence staining and western blot analysis. The present study identified that 10 µM of Zn supplementation prevented EMT changes, such as the loss of E-cadherin and the increase in α-smooth muscle actin and vimentin expression. Conversely, depletion of Zn with N,N,N',N'-tetrakis (2-pyridylmethyl)ethylenediamine in these cells aggravated changes in HG-induced EMT markers. Additionally, 10 µM Zn supplementation inhibited HG-induced transforming growth factor-ß1 overexpression and reactive oxygen species production. Of note, HG increased phosphatidylinositol 3-kinase/Akt (PI3K/Akt) and mitogen-activated protein kinase (MAPK) pathways activation and Zn reversed HG-induced expression of PI3K/Akt, extracellular-signal-regulated kinase (ERK) and p38 MAPK, as well as EMT proteins. Finally, inhibitors of PI3K/Akt, ERK and p38 MAPK, and Zn supplementation blocked the HG-induced EMT in NRK-52E cells. These results indicate that physiologically optimal levels of Zn can inhibit HG-induced EMT of the NRK-52E cells possibly through several mechanisms, including abrogation of HG-induced oxidative stress, and PI3K/Akt, p38 MAPK and ERK activation in NRK-52E cells.

Curcumin protects renal tubular epithelial cells from high glucose-induced epithelial-to-mesenchymal transition through Nrf2-mediated upregulation of heme oxygenase-1.

Curcumin has been observed to exhibit an anti-fibrotic effect in the liver, lung and gallbladder. However, the mechanisms underlying the cytoprotective effects of curcumin remain to be elucidated. The epithelial-to-mesenchymal transition (EMT) of mature tubular epithelial cells in the kidney is considered to contribute to the renal accumulation of matrix proteins associated with diabetic nephropathy. The EMT is also closely associated with the progression of renal interstitial fibrosis and oxidative stress. This process may occur through abrogation of high glucose (HG)-induced oxidative stress via activation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and heme oxygenase-1 (HO-1) in kidney tubular epithelial cells. In the present study, the effect of curcumin on HG-induced EMT in the NRK-52E normal rat kidney tubular epithelial cell line was investigated, and whether the effect of curcumin was mediated by the induction of Nrf2 and HO-1 expression was examined. The present study revealed that curcumin was able to prevent events associated with EMT, including the downregulation of E-cadherin and the increased expression of α-smooth muscle actin. Further analysis revealed that the expression levels of Nrf2 and HO-1 protein were elevated to a greater extent in the curcumin pretreated NRK-52E cells compared with those of the control. Notably, knockdown of Nrf2 with small interfering RNA prevented the curcumin-induced elevation in expression of HO-1 and the associated anti-fibrotic effects. In conclusion, the present findings suggested that curcumin may be significant in cellular antioxidant defense, through the activation of Nrf2 and HO-1, thereby protecting the NRK-52E cells from HG-induced EMT.

Zinc modulates high glucose-induced apoptosis by suppressing oxidative stress in renal tubular epithelial cells.

Hyperglycemia is a characteristic of diabetic nephropathy, inducing renal tubular cell apoptosis by eliciting oxidative stress and inflammation. Zinc (Zn) is known as an essential trace element in many enzymes and proteins involved in antioxidant defenses, electron transport, and exerting antiapoptotic or cytoprotective effects. In this study, the underlying mechanisms involved in the protective effects of Zn on high glucose-induced cytotoxicity were explored using cultured renal tubular epithelial cells (NRK-52E). The authors discovered that Zn supplementation inhibited high glucose (HG)-induced NRK-52E cell apoptosis by attenuating reactive oxygen species production, inhibiting HG-induced caspase-3 and caspase-9 activation, and inhibiting the release of cytochrome c from mitochondria to the cytosol. Further analysis revealed that Zn supplementation facilitated cell survival through increasing nuclear translocation of NF-E2-related factor 2 (Nrf2), leading to increased regulation of levels of two antioxidant enzymes, hemeoxygenase-1 and glutamate cysteine ligase, which provided an adaptive survival response against the HG-induced oxidative cytotoxicity. Moreover, the Zn-mediated increases in Nrf2 activity were suppressed by the pharmacological inhibition of Akt or extracellular signal-regulated kinase 1/2. Taken together, these findings suggest that Zn antiapoptosis capacity through the activation of Akt and ERK signal pathways leads to Nrf2 activation and, subsequently, Nrf2 target gene induction, thereby protecting the NRK-52E cells from HG-induced apoptosis.

Distribution of zinc and zinc transporters in the mouse ovarian follicles and corpus luteum.

Zinc is essential for female reproduction and it plays a role in sexual development, ovulation, menstruation and estrous cycles. Zinc deficiency may lead to female reproductive system dysfunction. The present study aimed to investigate the expression and distribution patterns of free zinc and the members of zinc transporter (ZnT) family, with zinc autometallographic (AMG), immunohistochemistry and real-time PCR, to explore the relationship of zinc homeostasis in the development and function of the ovary in the mouse. Our data revealed that the free zinc ions and ZnTs are predominantly distributed in the mouse ovarian follicles and corpus luteum. Specifically, AMG staining presented in various stages of the ovarian follicles and corpus luteum. ZnT1-9 mRNA was variously expressed, whereas ZnT10 mRNA was almost undetectable in the ovary. Moreover, the immunoreactivity of all the tested ZnTs, except for ZnT10, was detected with various intensity in the mouse primordial follicles, primary follicles, secondary follicles and antral follicles. In the corpus luteum, the immunoreactivity of ZnT1-5, 7, 8, 10, was abundantly observed in the granular and theca lutein cells and interstitial cells. Collectively, our results suggest that ZnT family proteins are differently distributed and might exert different biological functions in controlling cellular zinc levels, which regulate ovarian development and function in the mouse ovary.

Zinc transporter 5 and zinc transporter 7 induced by high glucose protects peritoneal mesothelial cells from undergoing apoptosis.

Zinc is an essential micronutrient and cytoprotectant involved in many types of apoptosis. The zinc transporter family SLC30A (ZnTs) is an important factor in the regulation of zinc homeostasis; however, its function in apoptosis in peritoneal mesothelial cells (PMCs) remains unknown. This study explores the regulation of zinc transporters and how they play a role in cell survival, particularly in rat peritoneal mesothelial cells (RPMCs), surrounding glucose concentrations, and the molecular mechanism involved. The messenger RNA (mRNA) transcripts were quantitatively measured by real-time polymerase chain reaction for all known nine zinc transport exporters (SLC30A1-8,10), as well as in primary RPMCs and the cells cultured under nonstimulated and HG-stimulated conditions. While many zinc transporters were constitutively expressed, ZnT5 mRNA and ZnT7 mRNA were strongly induced by HG. Overexpression of ZnT5 and ZnT7 respectively resulted in a decrease in the expression of caspace 3, caspace 8, BAX, and AIF and coincided with cell survival in the presence of HG. Inhibition of ZnT5 and ZnT7 expression using considerable siRNA-mediated knockdown of RPMCs was examined and, afterwards, the impact on cell apoptosis was investigated. Increased levels of apoptosis were observed after knockdown of ZnT5 and ZnT7. Furthermore, overexpression of ZnT5 and ZnT7 is accompanied by activation of PI3K/Akt pathway and inhibiting HG-induced apoptosis. This study suggests that the zinc transporting system in RPMCs is influenced by exposure to HG, particularly ZnT5 and ZnT7. This may account for the inhibition of HG-induced RPMC apoptosis and peritoneum injury, likely through targeting PI3K/Akt pathway-mediated cell survival.

Zinc transporter 7 induced by high glucose attenuates epithelial-to-mesenchymal transition of peritoneal mesothelial cells.

Zinc (Zn) is an essential micronutrient and cytoprotectant involved in preventing many types of epithelial-to-mesenchymal transition (EMT)-driven fibrosis in vivo. The zinc-transporter family SLC30A (ZnT) is a pivotal factor in the regulation of Zn homeostasis. However, its function in EMT in peritoneal mesothelial cells (PMCs) remains unknown. This study explored the regulation of zinc transporters and the role they play in cell EMT, particularly in rat peritoneal mesothelial cells (RPMCs), surrounding glucose concentrations and the molecular mechanism involved. The effects of high glucose (HG) on zinc transporter gene expression were measured in RPMCs by real-time PCR. We explored ZnT7 (Slc30A7): the effect of ZnT7 over-expression and siRNA-mediated knock-down on HG-induced EMT was investigated as well as the underlying molecular mechanisms. Over-expression of ZnT7 resulted in significantly inhibited HG-induced EMT in RPMCs, while inhibition of ZnT7 expression using a considerable siRNA-mediated knock-down of RPMCs increased the levels of EMT. Furthermore, over-expression of ZnT7 is accompanied by down-regulation of TGF-ß/Smad pathway, phospho-Smad3,4 expression levels. The finding suggests that the zinc-transporting system in RPMCs is influenced by the exposure to HG. The ZnT7 may account for the inhibition of HG-induced EMT in RPMCs, likely through targeting TGF-ß/Smad signaling.

Widespread expression of zinc transporter ZnT (SLC30) family members in mouse endocrine cells.

Zinc is abundant in most endocrine cell types, and plays a pivotal role in the synthesis and secretion of many hormones. Recent studies have demonstrated the expression of numerous zinc transporter (ZnT) family members in the pancreas, thyroid, and adrenal glands, suggesting a role for ZnTs in regulating cellular zinc homeostasis in endocrine cells. However, the cellular distribution of ZnTs in the endocrine organs has not been well established. In the present study, the mRNA expression level, cellular distribution of ZnTs as well as liable zinc ions were examined in the mouse pituitary, adrenal glands, thyroid, and pancreas. In general, ZnT1-10 mRNA was expressed to various degrees in the detected endocrine organs, with no detectable ZnT10 mRNA in the pancreas. In the anterior pituitary, both the acidophilic and basophilic cells were immunopositive to ZnT1-5, 7, 8, except for ZnT10. In the adrenal cortex, the immunoreactivity of all the tested ZnTs, including ZnT1-5, 7, 8, 10, was observed in the zona fasciculata, and some ZnTs were detected in the zona glomerulosa, zona reticularis, and the adrenal medulla. Both the follicle epithelial cells and parafollicular cells in the thyroid gland were immunostained with ZnT1-5, 7, 8, but not ZnT10. In the endocrine pancreas, the immunoreactivity of tested ZnTs was observed to various degrees except for ZnT10 in the cytoplasm of islet cells. Furthermore, autometallographic staining showed that liable zinc was observed in the endocrine cells, such as the adrenal cortical cells, thyroid follicle epithelial cells, and the pancreatic islet cells. All together, the wide distribution of liable zinc and the phenomenon that numerous ZnT family members are partially overlapped in a subset of endocrine cells suggest an important role for the ZnT family in controlling cellular zinc levels and subsequently regulating the synthesis and secretion of hormones in the endocrine system.

[Phase II clinical trial of sorafenib plus local chemotherapy in the treatment of metastatic renal cell carcinoma with pleural effusion].

OBJECTIVE: To evaluate the safety and efficacy of sorafenib plus cisplatin in the treatment of metastatic renal cell carcinoma (mRCC) with pleural effusion. METHODS: A total of 30 patients with mRCC (clear cell carcinoma) with pleural effusion from April 2009 to January 2011 were recruited. All received sorafenib 400 mg twice daily. And 11 patients in chemotherapy group received sorafenib plus local chemotherapeutic perfusion of cisplatin 40 mg weekly for 2 weeks while another 19 patients in control group received sorafenib alone. RESULTS: The response rate of pleural effusion was 10/11 for chemotherapy group versus 3/19 for control group (χ(2) = 13.097, P < 0.01). Followed up to April 30(th), 2011, 5 of 11 patients in chemotherapy group and 10 of 19 patients in control group died. Among those on sorafenib, the median overall survival time was 22 months (95%CI: 2.12 - 41.88) for local chemotherapy versus 9 months (95%CI: 8.20 - 9.80) without local therapy (P = 0.04). The most common events in local chemotherapy group were I-II thoracic pain, nausea and vomiting. And the incidence rates were 8/11 and 9/11 versus 4/19 and 3/19 respectively (P < 0.01). The main laboratory abnormalities were similar in two groups. CONCLUSION: The regimen of sorafenib plus pleural cavity perfusion of cisplatin is both effective and safe in the treatment of mRCC with pleural effusion. It may control local symptoms and achieve a better overall survival.

Disruption of the CaMKII/CREB signaling is associated with zinc deficiency-induced learning and memory impairments.

Many studies have shown that zinc deficiency not only retards growth, but also affects several brain functions, including learning and memory. However, the underlying mechanism of impaired hippocampus-dependent learning and memory under zinc deficiency is poorly understood. In this study, young mice were fed a zinc-deficient diet (0.85 ppm) for 5 weeks. Morris water maze result showed that zinc deficiency results in spatial learning impairment. We then examined whether zinc depletion-induced learning and memory defects are associated with changes in signaling molecules essential for the expression of long-term potentiation. Immunoblot results showed that the protein levels of calmodulin (CaM), phosphorylated CaM-dependent protein kinase II (CaMKII), and phosphorylated cAMP-responsive element binding protein (CREB) were significantly reduced, whereas the total protein levels of CaMKII and CREB did not change in the zinc-deficient hippocampus. Thus, we provide a previously unrecognized mechanism whereby zinc deficiency impairs hippocampal learning and memory, at least in part, through disruption of the CaM/CaMKII/CREB signaling pathway.