Mitochondrial folate pathway regulates myofibroblast differentiation and silica-induced pulmonary fibrosis.

BACKGROUND: Silica-induced pulmonary fibrosis (silicosis) is a diffuse interstitial fibrotic disease characterized by the massive deposition of extracellular matrix in lung tissue. Fibroblast to myofibroblast differentiation is crucial for the disease progression. Inhibiting myofibroblast differentiation may be an effective way for pulmonary fibrosis treatment. METHODS: The experiments were conducted in TGF-ß treated human lung fibroblasts to induce myofibroblast differentiation in vitro and silica treated mice to induce pulmonary fibrosis in vivo. RESULTS: By quantitative mass spectrometry, we revealed that proteins involved in mitochondrial folate metabolism were specifically upregulated during myofibroblast differentiation following TGF-ß stimulation. The expression level of proteins in mitochondrial folate pathway, MTHFD2 and SLC25A32, negatively regulated myofibroblast differentiation. Moreover, plasma folate concentration was significantly reduced in patients and mice with silicosis. Folate supplementation elevated the expression of MTHFD2 and SLC25A32, alleviated oxidative stress and effectively suppressed myofibroblast differentiation and silica-induced pulmonary fibrosis in mice. CONCLUSION: Our study suggests that mitochondrial folate pathway regulates myofibroblast differentiation and could serve as a potential target for ameliorating silica-induced pulmonary fibrosis.

Angiotensin 1-7 and its analogue decrease blood pressure but aggravate renal damage in preeclamptic mice.

Preeclampsia (PE) is a multisystem disease that affects the health of both the pregnant women and the fetus during pregnancy. Agonistic autoantibodies to the angiotensin II type I receptor (AT1-AA) play a significant role in the pathogenesis of PE. This study aimed to determine the effects of Angiotensin 1-7 (Ang 1-7) and its analogue AVE0991 on AT1-AA-induced PE model. Pregnant mice were divided into five groups: the normal pregnant group, AT1-AA-induced preeclampsia group, and AT1-AA-induced preeclampsia group treated with Losartan, Ang 1-7, and AVE0991, respectively. AT1-AA-induced PE model was established on gestational day 13 by tail intravenous injection of purified AT1-AA polyclonal antibody from serum of guinea pigs. Blood urea nitrogen (BUN), urine albumin and urinary creatinine were measured on day 18 of pregnancy. The systolic blood pressure (SBP) was measured from gestational day 13 to day 18. Renal structure changes were observed via light and electron microscopy. Compared with the normal pregnant group (NP group), AT1-AA-induced preeclampsia group (PE group) exhibited elevated blood pressure and proteinuria, consistent with the characteristics of PE. Ang 1-7 or AVE0991 treatment decreased blood pressure without showing renoprotective effects. The findings indicated that Ang 1-7 and its analogue reduced blood pressure but aggravated renal damage in AT1-AA-induced PE mice.

How different online self-presentations relate to life satisfaction differently in college students: The role of positive online feedback and self-esteem.

In the Internet age, some online factors, such as online self-presentation, related to life satisfaction have received much attention. However, it is unclear whether and how different strategies of online self-presentation are linked to an individual's life satisfaction differently. Accordingly, the present study examined the possible different relationships between different online self-presentations and life satisfaction with a sample of 460 Chinese college students. Using a series of questionnaires, a moderated mediation model was built in which positive online feedback was a mediator and self-esteem was a moderator. The results indicated that: (1) positive self-presentation was negatively associated with college students' life satisfaction, whereas honest self-presentation was positively related to it; (2) positive online feedback was a significant mediator in such relationships; (3) the mediation process was moderated by self-esteem. Specifically, positive self-presentation was negatively related to positive online feedback only for high self-esteem college students, but negatively associated with life satisfaction only for low self-esteem ones. By contrast, honest self-presentation was positively associated with positive online feedback despite the level of self-esteem, but positively linked with life satisfaction only for those with low self-esteem. The findings suggest that honest rather than positive online self-presentation should be conducive to college students' life satisfaction, particularly for those with low self-esteem. The implications were discussed.

Empagliflozin Ameliorates Preeclampsia and Reduces Postpartum Susceptibility to Adriamycin in a Mouse Model Induced by Angiotensin Receptor Agonistic Autoantibodies.

Preeclampsia (PE) is the leading cause of maternal and perinatal morbidity and mortality and also is a risk factor for cardiovascular and kidney disease later in life. PE is associated with oversecretion of autoantibodies against angiotensin II type 1 receptor (AT1-AA) by the placenta into the maternal circulation. Here, we sought to determine the therapeutic value of the sodium-glucose co-transporter 2 (SGLT2) inhibitor empagliflozin (EMPA) in mice with AT1-AA-induced preeclampsia. Pregnant mice were injected with AT1-AA at gestation day (GD) 13 and treated daily with EMPA until GD 19, at which point some of the maternal mice were sacrificed and assessed. The other maternal mice were labored on time and challenged with adriamycin (ADR) at 12 weeks postpartum; their offspring were assessed for fetal outcomes. We showed that EMPA treatment significantly relieved high systolic blood pressure and proteinuria and ameliorated kidney injury in PE mice without affecting fetal outcomes. EMPA also ameliorated podocyte injury and oxidative stress, reduced the expression of SGLT2 and activated the AMPK/SIRT1 signaling pathway in vivo and in vitro. Remarkably, EMPA treatment during pregnancy reduced ADR-induced kidney and podocyte injury postpartum. These findings suggest that EMPA could be a potential pharmacological agent for PE.

TGF-ß promote epithelial-mesenchymal transition via NF-κB/NOX4/ROS signal pathway in lung cancer cells.

Epithelial-mesenchymal transition (EMT), transforming growth factor ß(TGF-ß) and reactive oxygen species(ROS) plays a central role in cancer metastasis. Moreover, nicotinamide adenine dinucleotide phosphate 4(NOX4) is one of the main sources of ROS in lung cancer cells suggesting that NOX4 is associated with tumor cell migration. NF-κB(Nuclear factor-Kappa-B) is known to regulate ROS-mediated EMT process by activating Snail transcription factor in A549 cells. The purpose of this study was to explore the relationship between NF-κB and NOX4 in ROS production during TGF-ß induced EMT process. Several fractions have been pooled to evaluates the EMT process on lung cancer cells through real-time PCR, Western Blot and flow cytometry with DCFH-DA probe etc. Cells proliferation and migration activities were monitored by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay and wound healing assay respectively. The result showed that TGF-ß induction decreased the expression of E-cadherin, increased the Vimentin and the EMT transcription factor Snail in A549 cells. DPI (Diphenyleneiodonium chloride, an inhibitor of NOX4) inhibited the NOX4 expression and reduced ROS production induced by TGF-ß, but didn't affect the activation of NF-κB induced by TGF-ß (P > 0.05). BAY11-7082 (an inhibitor of NF-κB) inhibited the NF-κB (p65) expression and prevented the increase of NOX4 expression and ROS production induced by TGF-ß (P < 0.001), which has also verified reduced TGF-ß induced cell migration by inhibiting the EMT process, and also reduced cell proliferation of A549 cells (P < 0.001). The current research confirmed the TGF-ß mediated EMT process via NF-κB/NOX4/ROS signaling pathway, NF-κB and NOX4 are likely to be the potential therapeutic targets for lung cancer metastasis.

Overexpression of heat shock protein 70 inhibits epithelial-mesenchymal transition and cell migration induced by transforming growth factor-ß in A549 cells.

Heat shock protein 70 (HSP70) is a key member of the HSP family that contributes to a pre-cancerous environment; however, its role in lung cancer remains poorly understood. The present study used geranylgeranylacetone (GGA) to induce HSP70 expression, and transforming growth factor-ß (TGF-ß) was used to construct an epithelial-mesenchymal transition (EMT) model by stimulating A549 cells in vitro. Western Blot was performed to detect protein levels of NADPH oxidase 4 (NOX4) and the EMT-associated proteins E-cadherin and vimentin both before and after HSP70 expression. Cell morphological changes were observed, and the effect of HSP70 on cell migration ability was detected via the wound healing. The results demonstrated that GGA at 50 and 200 µmol/L could significantly induce HSP70 expression in A549 cells (P < 0.05). Furthermore, HSP70 induced by 200 µmol/L GGA significantly inhibited the changes of E-cadherin, vimentin, and cell morphology induced by TGF-ß (P < 0.05), while HSP70 induced by 50 µmol/L GGA did not. The results of the wound healing assay indicated that 200 µmol/L GGA significantly inhibited A549 cell migration induced by TGF-ß. Taken together, the results of the present study demonstrated that overexpression of HSP70 inhibited the TGF-ß induced EMT process and changed the cell morphology and migratory ability induced by TGF-ß in A549 cells.

Folate and macrophage folate receptor-ß in idiopathic pulmonary fibrosis disease: the potential therapeutic target?

Idiopathic pulmonary fibrosis (IPF) is a chronic, fatal disease with high mortality and poor prognosis. It is characterized by a gradual decline in lung function, and there are currently no effective therapeutic methods. Folate is a water-soluble B vitamin that plays an important role in one-carbon transfer reactions, nucleic acid biosynthesis and methylation reactions. Studies have shown that folate may participate in the pathogenesis of IPF through ways of DNA repair, methylation, and reactive oxygen species. Macrophage activation is an important early cellular event in IPF and the inflammatory response that they trigger is a significant feature of IPF. Folate receptor-ß (FR-ß) is a cell surface glycosylphosphatidylinositol-anchored glycoprotein that can mediate the unidirectional transport of folate into cells. And it has been found in previous studies that FR-ß is usually overexpressed on activated macrophages, but the expression on resting macrophages was undetectable. Therefore, targeting FR-ß may have potential value for the early diagnosis and therapy of IPF. Our goal is to highlight the biological role of folate and FR-ß in IPF, and we hope to provide helpful insight for clinical treatment strategies.

Panax Notoginseng Ameliorates Podocyte EMT by Targeting the Wnt/ß-Catenin Signaling Pathway in STZ-Induced Diabetic Rats.

INTRODUCTION: Epithelial-mesenchymal transition (EMT) may contribute to podocyte dysfunction in diabetic nephropathy (DN). Aiming to identify novel therapeutic options, we investigated the protective effects of Panax notoginseng (PN) on podocyte EMT in diabetic rats and explored its mechanisms. METHODS: Diabetes was induced in rats with streptozotocin (STZ) by intraperitoneal injection at 55 mg/kg. Diabetic rats were randomly divided into three groups, namely, diabetic rats, diabetic rats treated with beraprost sodium (BPS) at 0.6 mg/kg/d or PN at 0.4 g/kg/d p.o., for 12 weeks. Urinary albumin/creatinine ratio (ACR), biochemical parameters, renal histopathology, and podocyte morphological changes were evaluated. Protein expression of EMT markers (desmin, α-SMA, and nephrin) as well as components of the Wnt/ß-catenin pathway (wnt1, ß-catenin, and snail) was detected by immunohistochemistry and Western blot, respectively. RESULTS: In diabetic rats, severe hyperglycemia and albuminuria were detected. Moreover, mesangial expansion and podocyte foot process effacement were found markedly increased in diabetic kidneys. Increased protein expression of wnt1, ß-catenin, snail, desmin, and α-SMA, as well as decreased protein expression of nephrin was detected in diabetic kidneys. All these abnormalities found in DN rats were partially restored by PN treatment. CONCLUSION: PN ameliorated albuminuria and podocyte EMT in diabetic rats partly through inhibiting Wnt/ß-catenin signaling pathway. These findings provide experimental arguments for a novel therapeutic option in DN.

Xuesaitong Protects Podocytes from Apoptosis in Diabetic Rats through Modulating PTEN-PDK1-Akt-mTOR Pathway.

Diabetic kidney disease (DKD) is a major cause of end-stage renal disease (ESRD), and therapeutic strategies for delaying its progression are limited. Loss of podocytes by apoptosis characterizes the early stages of DKD. To identify novel therapeutic options, we investigated the effects of Xuesaitong (XST), consisting of total saponins from Panax notoginseng, on podocyte apoptosis in streptozotocin- (STZ-) induced diabetic rats. XST (5 mg/kg·d) or Losartan (10 mg/kg·d) was given to diabetic rats for 12 weeks. Albuminuria, renal function markers, and renal histopathology morphological changes were examined. Podocyte apoptosis was determined by triple immunofluorescence labelling including a TUNEL assay, WT1, and DAPI. Renal expression of Nox4, miRNA-214, PTEN, PDK1, phosphorylated Akt, mTOR, and mTORC1 was detected. In diabetic rats, severe hyperglycaemia and albuminuria developed, and apoptotic podocytes were markedly increased in diabetic kidneys. However, XST attenuated albuminuria, mesangial expansion, podocyte apoptosis, and morphological changes of podocytes in diabetic rats. Decreased expression of PTEN, as well as increased expression of Nox4, miRNA-214, PDK1, phosphorylated Akt, mTOR, and mTORC1, was detected. These abnormalities were partially restored by XST treatment. Thus, XST ameliorated podocyte apoptosis partly through modulating the PTEN-PDK1-Akt-mTOR pathway. These novel findings might point the way to a natural therapeutic strategy for treating DKD.

HSP70 regulates cell proliferation and apoptosis in actinomycin-D-treated lung cancer cells.

BACKGROUND: Heat-shock protein 70 (HSP70) is a member of the heat-shock protein family which is expressed in various types of cancer and associated with apoptosis in cancer cells. However, the role of HSP70 in the regulation of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38MAPK)-dependent growth and apoptosis in lung cancer cells remains largely unknown. METHODS: In this study, we conducted in vitro experiments. First, we examined the effect of HSP70 by treatment with mild heat and JNK/p38 MAPK inhibitors on cell proliferation in A549 cells by MTT assay. And then, through the use of flow cytometric assay, we examined the effect of HSP70 by treatment with mild heat and JNK/p38 MAPK inhibitors on cell apoptosis in A549 cells. Finally, we determined the role of HSP70 in the regulation of JNK and p38 MAPK-dependent growth and apoptosis in A549 cells by siRNA HSPAIA-2009 and Western blot. RESULTS: We found that treatment of the cells with mild heat and JNK/p38 MAPK pathway inhibitors (SP600125 and SB203580) promoted cell proliferation in the presence of actinomycin D (ActD) in A549 cells. We also showed that treatment with mild heat or SP600125 and SB203580 significantly slowed the steps of apoptosis induced by ActD in A549 cells. Moreover, we found that HSP70 overexpression induced by mild heat markedly decreased the expression of cell growth and apoptosis-related protein p-JNK, p38 and caspase-3. By contrast, knockdown of HSP70 by siRNA HSPAIA-2009 effectively promoted the expression of the JNK and p38 MAPK. CONCLUSIONS: Our results indicate that HSP70 plays an important role in lung cancer growth and apoptosis through the activation of JNK and p38 MAPK signaling in actinomycin-D-treated lung cancer A549 cells.

Effects of cyclophosphamide on the phenotypes and functions of THP-1 cells.

Early and accurate evaluation of immunotoxicity is crucial. However, there are few in vitro models for immunosuppressive evaluation. THP-1 cells has long been used for in vitro sensitivity evaluation. Whether it can be used for immunosuppressive evaluation remains unclear. In this study, effects of immunosuppressant cyclophosphamide (CY) on THP-1 cells were observed while 2, 4-Dinitrochlorobenzene (DNCB) was used as a control. The phenotypes of THP-1 cells, the ability to activate naïve T cells, intracellular reactive oxygen species (ROS) level, gene markers, phagocytic ability and cell apoptosis were detected after THP-1 cells being exposed to different concentrations of CY and DNCB. Both CY and DNCB were able to activate THP-1 cells, but there were a lot of differences in their effects on THP-1 cells, such as the changes in phenotypes, in the ability to activate naïve T cells, in ROS production and in marker gene expression. Firstly, CY down-regulated the expression of CD86 on THP-1 cells while DNCB up-regulated its expression. Secondly, the ability of THP-1 cells to activate naïve T cells was enhanced by CY and suppressed by DNCB. Thirdly, CY raised rapid and transient elevation of ROS level in THP-1 cells, while the effects of DNCB were slower and longer-lasting. Finally, only CY could lead to an increase in heme oxygenase 1 (HMOX1) expression. Taken all these results into account, we suggested that THP-1 cell line possesses the potency to be an in vitro model of immunosuppressive evaluation. And the surface molecule CD86, the ability to activate naïve T cells, the ROS production and the gene marker HMOX1 of THP-1 cells are promising markers.

Astragalus membranaceus and Panax notoginseng, the Novel Renoprotective Compound, Synergistically Protect against Podocyte Injury in Streptozotocin-Induced Diabetic Rats.

This study was aimed at investigating the synergistical protective effects of Astragalus membranaceus (AG) and Panax notoginseng (NG) on podocyte injury in diabetic rats. Diabetes was induced in rats by a single intraperitoneal injection of streptozotocin at 55 mg/kg. Diabetic rats were then orally administrated with losartan, AG, NG, and AG plus NG (2 : 1) for 12 weeks. Albuminuria, biochemical markers, renal histopathology, and podocyte number per glomerulus were measured. Podocyte apoptosis was determined by triple immunofluorescence labeling including TUNEL assay, WT1, and DAPI. Renal expression of nephrin, α-dystroglycan, Bax, Bcl-xl, and Nox4 was evaluated by immunohistochemistry, western blot, and RT-PCR. AG plus NG ameliorated albuminuria, renal histopathology, and podocyte foot process effacement to a greater degree than did AG or NG alone. The number of podocytes per glomerulus, as well as renal expression of nephrin, α-dystroglycan, and Bcl-xl, was decreased, while podocyte apoptosis, as well as renal expression of Bax and Nox4, was increased in diabetic rats. All of these abnormalities were partially restored by AG plus NG to a greater degree than did AG or NG alone. In conclusion, AG and NG synergistically ameliorated diabetic podocyte injury partly through upregulation of nephrin, α-dystroglycan, and Bcl-xl, as well as downregulation of Bax and Nox4. These findings might provide a novel treatment combination for DN.

miR-let-7d attenuates EMT by targeting HMGA2 in silica-induced pulmonary fibrosis.

Silicosis is a serious occupational disease characterized by pulmonary chronic inflammation and progressive fibrosis. Epithelial-mesenchymal transition (EMT) of alveolar epithelial cells plays a vital role in silicosis. Recent studies discovered a variety of microRNAs (miRNAs) participating in fibrotic diseases. Here, we aimed to explore the function and mechanism of miRNA let-7d in the EMT process in silica-induced alveolar epithelial cells. To detect whether let-7d and its target HMGA2 were involved in silica-induced EMT, we established a silicosis mouse model and found that let-7d was down-regulated and HMGA2 was up-regulated in the silica-treated group. Then we applied an in vitro co-culture system to imitate the EMT process in A549 cells after silica treatment. The down-regulation of let-7d and up-regulation of HMGA2 were also observed in vitro. The knockdown of HMGA2 significantly inhibited the silica-induced EMT. Furthermore, we found that overexpression of let-7d could reduce the expression of HMGA2 and consequently inhibited the silica-induced EMT, whereas inhibition of let-7d increased the expression of HMGA2 and promoted the silica-induced EMT. In conclusion, let-7d negatively regulated silica-induced EMT and inhibited silica-induced pulmonary fibrosis, which might be partially realized by directly binding to HMGA2. Our data suggested that miRNA let-7d might have a potential protective effect in the fibrotic process and become a new therapeutic target for silicosis or other fibrotic diseases.

Downregulation of exosomal let-7a-5p in dust exposed- workers contributes to lung cancer development.

BACKGROUND: Either chronic or acute exposure to dust particles may lead to pneumoconiosis and lung cancer, and lung cancer mortality among patients diagnosed with pneumoconiosis is increasing. Utilizing genome-wide sequencing technology, this study aimed to identify methods to decrease the number of patients with pneumoconiosis who die from lung cancer. METHODS: One hundred fifty-four subjects were recruited, including 54 pneumoconiosis patients and 100 healthy controls. Exosomes were isolated from the venous blood of every subject. Distinctive miRNAs were identified using high throughput sequencing technology, and bioinformatics analysis predicted target genes involved in lung cancer as well as their corresponding biological functions. Moreover, cross-cancer alterations of genes related to lung cancer were investigated, and survival analysis was performed using 2437 samples with an average follow-up period of 49 months. RESULTS: Let-7a-5p was revealed to be downregulated by 21.67% in pneumoconiosis. Out of the 683 let-7a-5p target genes identified from bioinformatics analysis, four genes related to five signaling pathways were confirmed to be involved in lung cancer development. Alterations in these four target genes were then explored in 4105 lung cancer patients, and BCL2L1 and IGF1R were demonstrated to be aberrantly expressed. Survival analysis further revealed that high expression of BCL2L1 corresponded to reduced survival of lung cancer patients (HR (95%CI) = 1.75(1.33~2.30)), while patient survival time was unaffected by expression of IGF1R (HR (95%CI) = 1.15 (0.98~1.36)). CONCLUSIONS: In patients with lung adenocarcinoma, simultaneous downregulation of exosomal let-7a-5p and elevated expression of BCL2L1 are useful as predictive biomarkers for poor survival.

Mechanistic Insight and Management of Diabetic Nephropathy: Recent Progress and Future Perspective.

Diabetic nephropathy (DN) is the most serious microvascular complication of diabetes and the largest single cause of end-stage renal disease (ESRD) in many developed countries. DN is also associated with an increased cardiovascular mortality. It occurs as a result of interaction between both genetic and environmental factors. Hyperglycemia, hypertension, and genetic predisposition are the major risk factors. However, the exact mechanisms of DN are unclear. Despite the benefits derived from strict control of glucose and blood pressure, as well as inhibition of renin-angiotensin-aldosterone system, many patients continue to enter into ESRD. Thus, there is urgent need for improving mechanistic understanding of DN and then developing new and effective therapeutic approaches to delay the progression of DN. This review focuses on recent progress and future perspective about mechanistic insight and management of DN. Some preclinical relevant studies are highlighted and new perspectives of traditional Chinese medicine (TCM) for delaying DN progression are discussed in detail. These findings strengthen the therapeutic rationale for TCM in the treatment of DN and also provide new insights into the development of novel drugs for the prevention of DN. However, feasibility and safety of these therapeutic approaches and the clinical applicability of TCM in human DN need to be further investigated.