ABSTRACT
Sex differences in kidney stone formation are well known. Females generally have slightly acidic blood and higher urine pH when compared with males, which makes them more vulnerable to calcium stone formation, yet the mechanism is still unclear. We aimed to examine the role of sex in stone formation during hypercalciuria and urine alkalinization through acetazolamide and calcium gluconate supplementation, respectively, for 4 weeks in wild-type (WT) and moderately hypercalciuric [TRPC3 knockout [KO](-/-)] male and female mice. Our goal was to develop calcium phosphate (CaP) and CaP+ calcium oxalate mixed stones in our animal model to understand the underlying sex-based mechanism of calcium nephrolithiasis. Our results from the analyses of mice urine, serum, and kidney tissues show that female mice (WT and KO) produce more urinary CaP crystals, higher [Ca2+], and pH in urine compared to their male counterparts. We identified a sex-based relationship of stone-forming phenotypes (types of stones) in our mice model following urine alkalization/calcium supplementation, and our findings suggest that female mice are more susceptible to CaP stones under those conditions. Calcification and fibrotic and inflammatory markers were elevated in treated female mice compared with their male counterparts, and more so in TRPC3 KO mice compared with their WT counterparts. Together these findings contribute to a mechanistic understanding of sex-influenced CaP and mixed stone formation that can be used as a basis for determining the factors in sex-related clinical studies.
Subject(s)
Hypercalciuria , Kidney Calculi , Mice, Knockout , Phenotype , Animals , Female , Male , Hypercalciuria/metabolism , Hypercalciuria/urine , Mice , Kidney Calculi/metabolism , Kidney Calculi/urine , Kidney Calculi/etiology , Calcium Phosphates/metabolism , Calcium Phosphates/urine , Hydrogen-Ion Concentration , Mice, Inbred C57BL , Disease Models, Animal , Kidney/metabolism , Sex Factors , Sex Characteristics , Calcium Oxalate/metabolism , Calcium Oxalate/urine , TRPC Cation Channels/metabolism , TRPC Cation Channels/geneticsABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Qian Yang Yu Yin granules (QYYYG) have a long history in the treatment of hypertensive renal damage (HRD) in China. Clinical studies have found that QYYYG stabilizes blood pressure and prevents early renal damage. However, the exact mechanism is not entirely clear. AIM OF THE STUDY: To evaluate the therapeutic effect and further explore the therapeutic mechanism of QYYYG against HRD. MATERIALS AND METHODS: The efficacy of QYYYG in treating HRD was assessed in spontaneous hypertension rats (SHR). Renal autophagy and the TRPC6-CaMKKß-AMPK pathway in rats were evaluated. The regulatory role of QYYYG in angiotensin II (Ang II) induced abnormal autophagy in rat podocytes was determined by detecting autophagy-related proteins, intracellular Ca2+ content, and the TRPC6-CaMKKß-AMPK-mTOR pathway expressions. Finally, we established a stable rat podocyte cell line overexpressing TRPC6 and used the cells to verify the regulatory effects of QYYYG. RESULTS: QYYYG alleviated HRD and reversed the abnormal expression of autophagy-related genes in the SHR. In vitro, QYYYG protected against Ang II-induced podocyte damage. Furthermore, treatment of podocytes with QYYYG reversed Ang II-induced autophagy and inhibited Ang II-stimulated TRPC6 activation, Ca2+ influx and activation CaMKKß-AMPK pathway. Overexpression of TRPC6 resulted in pronounced activation of CaMKKß, AMPK, and autophagy induction in rat podocytes, which were significantly attenuated by QYYYG. CONCLUSIONS: The present study suggested that QYYYG may exert its HRD protective effects in part by regulating the abnormal autophagy of podocytes through the TRPC6-CaMKKß-AMPK-mTOR pathway.
Subject(s)
Hypertension , Podocytes , Animals , Rats , Calcium-Calmodulin-Dependent Protein Kinase Kinase/metabolism , TRPC6 Cation Channel/metabolism , AMP-Activated Protein Kinases/metabolism , Calcium/metabolism , Autophagy , TOR Serine-Threonine Kinases/metabolism , Angiotensin II/metabolism , Hypertension/drug therapy , Hypertension/metabolism , TRPC Cation Channels/genetics , TRPC Cation Channels/metabolism , TRPC Cation Channels/pharmacologyABSTRACT
Tetrandrine (Tet), a compound found in a traditional Chinese medicine, presents the protective effect for kidney function. Our study is aimed at clarifying the efficacy and underlying mechanism of Tet on podocyte injury. In this study, podocyte injury was induced in rats with adriamycin (ADR), and MPC5 podocytes were constructed with TRPC6 overexpression. We found that Tet treatment reduced the levels of proteinuria, serum creatinine, and blood urea nitrogen and increased plasma albumin levels in ADR-induced rats. Tet reduced intracellular Ca2+ influx and apoptosis in MPC5 podocytes overexpressing TRPC6. Tet downregulated the expression of renal TRPC6, RhoA, and ROCK1 and upregulated the expression of synaptopodin; meanwhile, it reduced calcineurin activity in vivo and in vitro. In conclusion, Tet protects against podocyte by affecting TRPC6 and its downstream RhoA/ROCK1 signaling pathway.
Subject(s)
Podocytes , Animals , Benzylisoquinolines , Calcineurin/metabolism , Calcineurin/pharmacology , Creatinine , Doxorubicin/metabolism , Doxorubicin/pharmacology , Podocytes/metabolism , Rats , Serum Albumin/metabolism , Serum Albumin/pharmacology , TRPC Cation Channels/metabolism , TRPC Cation Channels/pharmacology , TRPC6 Cation Channel/metabolism , rho-Associated Kinases/metabolism , rho-Associated Kinases/pharmacology , rhoA GTP-Binding Protein/metabolismABSTRACT
Hypoxia-induced pulmonary arterial hypertension (HPAH) is due to hypoxia caused by vascular endothelial cell remolding and damage. Previous studies have suggested that CX3CL1 plays an important role in HPAH which is affected by oxidative stress. Ca2+ channel activation correlated with increasing NF-κB levels induced by ROS. Tanreqing injection (TRQ) is a traditional Chinese medicine (TCM) for acute upper respiratory tract infection and acute pneumonia. In the present study, we explored the effect of TRQ on human pulmonary artery smooth muscle cells (HPASMCs) undergoing hypoxia and feasible molecular mechanisms involved in. Cell proliferation was assayed using CCK8 kits. Immunofluorescence and western blotting along with ELISA assay were performed to investigate the effect of TRQ on hypoxia-induced ROS, Ca2+, hydroxyl free radicals, and the expression of Ca2+ channel protein TRPC1, CX3CR1, HIF-1α, NF-κBp65, and p-NF-κBp65 in HPASMCs. Human CX3CL1 and the inhibitor of TRPC1 as SKF96365 were used for further investigation. TRQ inhibited hypoxia-induced increasing cell adhesion, ROS, Ca2+, hydroxyl free radicals, CX3CR1, HIF-1α, NF-κBp65 activation, and even on TRPC1 expression in HPASMC which tended to be attenuated even reversed by CX3CL1. Our results suggested that TRQ might help to attenuate remodeling of HPASMC through inhibiting the ROS and TRPC1/CX3CL1 signaling pathway.
Subject(s)
Cell Hypoxia/drug effects , Chemokine CX3CL1/metabolism , Drugs, Chinese Herbal/therapeutic use , Hypertension, Pulmonary/drug therapy , Hypertension, Pulmonary/etiology , Myocytes, Smooth Muscle/drug effects , Pulmonary Artery/drug effects , TRPC Cation Channels/metabolism , Cell Proliferation , Drugs, Chinese Herbal/pharmacology , Humans , Hypertension, Pulmonary/physiopathologyABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Magnolia officinalis Cortex (M. officinalis) is a classical traditional Chinese medicine (TCM) widely used to treat digestive system diseases. It effectively regulates gastrointestinal motility to improve abdominal pain, abdominal distension and other symptoms. Magnolol (MAG) and honokiol (HON) are the main pharmacodynamic components responsible for the gastrointestinal activity of M. officinalis. AIM OF THE STUDY: The transient receptor potential (TRP) family is highly expressed in the gastrointestinal tract and participates in the regulation of gastrointestinal motility, visceral hypersensitivity, visceral secretion and other physiological activities. In this study, the calcium-lowering mechanisms of MAG and HON contributing to the smooth muscle relaxation associated with TRP are discussed. MATERIALS AND METHODS: The relaxation smooth muscle effects of MAG and HON were tested by the isolated intestine tone tests. A synthetic MAG probe (MAG-P) was used to target fishing for their possible target. The distribution of MAG on the smooth muscle was identified by a molecular tracer based on chemical biology. Ca2+ imaging and dual-luciferase reporter assays were used to determine the effects on the target proteins. Finally, the calcium-mediating effects of MAG and HON on smooth muscle cells and TRPC4-knockdown cells were compared to verify the potential mechanism. RESULTS: After confirming the smooth muscle relaxation in the small intestine induced by MAG and HON, the relaxation effect was identified mainly due to the downregulation of intracellular calcium by controlling external calcium influx. Although MAG and HON inhibited both TRPV4 and TRPC4 channels to reduce calcium levels, the inhibitory effect on TRPC4 channels is an important mechanism of their smooth muscle relaxation effect, since TRPC4 is widely expressed in the small intestinal smooth muscle cells. CONCLUSIONS: The inhibition of MAG and HON on TRPC4 channels contributes to the relaxation of intestinal smooth muscle.
Subject(s)
Biphenyl Compounds/pharmacology , Calcium Signaling/drug effects , Intestines/drug effects , Lignans/pharmacology , Muscle, Smooth/drug effects , Animals , HEK293 Cells , Humans , Male , Medicine, Chinese Traditional , Muscle Contraction/drug effects , Myocytes, Smooth Muscle/drug effects , Rats , Rats, Sprague-Dawley , TRPC Cation Channels/drug effects , TRPV Cation Channels/drug effectsABSTRACT
Aging is an inescapable complex physiological but extendable process, and all cells, including stem cells, are altered over time. Diverse mechanism(s) could modulate stem cell number, their proliferation rate, and promote tissue repair during aging that leads to longevity. However, the factors that could restore aging stem cell potency and would lead to healthy aging are not fully identified. Here we show that maintaining cytosolic Ca2+ levels was essential for modulating stem cells function in aged mesenchymal stem cells (MSCs). Increasing external Ca2+ induced spindle shape stem cell morphology and maintained stem cell surface marker expression in aged bone marrow-derived MSCs. Similarly, stem cell survival and proliferation of aged MSCs was dependent on cytosolic Ca2+ levels. Importantly, Ca2+ entry potentiated cell cycle progression, and stem cell potential was increased in cells incubated with higher external Ca2+. Moreover, blocking Ca2+ entry using SKF 96365, decreased stem cell survival and its proliferation but, treatment with 2-APB did not significantly affected cell proliferation, rather only modulated cell viability. Evaluation of Ca2+ entry channels, showed that TRPC1/Orai1/Orai3 and their regulator STIM1 was essential for MSCs proliferation/viability as gene silencing of Orai1/Orai3/TRPC1/STIM1 significantly inhibited stem cell viability. Finally, MSCs isolated from aged mice that were subjected to higher Ca2+ levels, were able to rescue age-induced loss of MSCs function. Together these results suggest that Ca2+ entry is essential for preventing the loss of aged stem cell function and supplementing Ca2+ not only restored their proliferative potential but, allowed them to develop into younger stem cell lineages that could be critical for regenerative medicine.
Subject(s)
Calcium Channels , Mesenchymal Stem Cells , Animals , Calcium/metabolism , Cell Proliferation , Cells, Cultured , Mesenchymal Stem Cells/metabolism , Mice , ORAI1 Protein , Stem Cells/metabolism , Stromal Interaction Molecule 1 , TRPC Cation ChannelsABSTRACT
BACKGROUND: Atractylodis rhizoma, an aromatic herb for resolving dampness, is used to treat Kidney-related edema in traditional Chinese medicine for thousands years. This herb possesses antioxidant effect. However, it is not yet clear how Atractylodis rhizoma prevents glomerular injury through its anti-oxidation. PURPOSE: Based the analysis of Atractylodis rhizoma water extract (ARE) components and network pharmacology, this study was to explore whether ARE prevented glomerular injury via its anti-oxidation to inhibit oxidative stress-driven transient receptor potential channel 6 (TRPC6) and its downstream molecule calcium/calmodulin-dependent protein kinase IV (CaMK4) signaling. METHODS: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to analyze ARE components. Network pharmacology analysis was preliminarily performed. Male Sprague-Dawley rats were given 10% fructose drinking water (100 mL/d) for 16 weeks. ARE at 720 and 1090 mg/kg was orally administered to rats for the last 8 weeks. Hydrogen peroxide (H2O2) and malondialdehyde (MDA) level, and superoxide dismutase (SOD) activity in rat kidney cortex were detected, respectively. In rat glomeruli, redox-related factors forkhead box O3 (FoxO3), SOD2 and catalase (CAT), podocyte slit diaphragm proteins podocin and nephrin, cytoskeleton proteins CD2-associated protein (CD2AP) and α-Actinin-4, as well as TRPC6, p-CaMK4 and synaptopodin protein levels were analyzed by Western Blotting. SOD2 and CAT mRNA levels were detected by qRT-PCR. RESULTS: 36 components were identified in ARE. Among them, network pharmacology analysis indicated that ARE might inhibit kidney oxidative stress. Accordingly, ARE up-regulated nuclear FoxO3 expression, and then increased SOD2 and CAT at mRNA and protein levels in glomeruli of fructose-fed rats. It reduced H2O2 and MDA levels, and increased SOD activity in renal cortex of fructose-fed rats. Subsequently, ARE down-regulated TRPC6 and p-CaMK4, and up-regulated synaptopodin in glomeruli of fructose-fed rats. Furthermore, ARE increased podocin and nephrin, as well as CD2AP and α-Actinin-4, being consistent with its reduction of urine albumin-to-creatinine ratio and improvement of glomerular structure injury in this animal model. CONCLUSIONS: These results suggest that ARE may prevent glomerular injury in fructose-fed rats possibly by reducing oxidative stress to inhibit TRPC6/p-CaMK4 signaling and up-regulate synaptopodin expression. Therefore, ARE may be a promising drug for treating high fructose-induced glomerular injury in clinic.
Subject(s)
Atractylodes , Calcium-Calmodulin-Dependent Protein Kinase Type 4/metabolism , Kidney Diseases/drug therapy , Plant Extracts/pharmacology , TRPC Cation Channels/metabolism , Animals , Atractylodes/chemistry , Chromatography, Liquid , Fructose/adverse effects , Hydrogen Peroxide/metabolism , Kidney/drug effects , Kidney Diseases/chemically induced , Male , Oxidation-Reduction , Oxidative Stress , Rats , Rats, Sprague-Dawley , Rhizome/chemistry , Signal Transduction , TRPC6 Cation Channel , Tandem Mass SpectrometryABSTRACT
OBJECTIVE: To determine the effect of Wenyang Huazhuo Fang (WHF), a Traditional Chinese Medicine decoction, on renal function in a rat model of doxorubicin-induced nephropathy, and to elucidate the underlying mechanism. METHODS: Sprague-Dawley rats were randomly divided into six groups: control, doxorubicin-nephropathy, and prednisone-treated (6.45 mg·kg-1·dï¼1) doxorubicin nephropathy groups, as well as high- (7.26 g·kg-1·dï¼1, medium- (2.42 g·kg-1·dï¼1, and low-dose (0.81 g·kg-1·dï¼1 WHF-treated doxorubicin-nephropathy groups. The nephropathy rat model was established by two tail vein injections of doxorubicin, followed by prednisone or WHF treatment for 8 weeks. Body weights were monitored and urinary protein was measured every 2 weeks. After the end of the treatment period, the rats were euthanized. Serum biochemical indicators were determined and renal morphological alterations were assessed using histological staining. The expression of transient receptor potential cation channel subfamily C member 6 (TRPC6), stromal interaction molecule 1 (STIM1), and calcium release-activated calcium channel protein 1 (Orai1) was detected using western blotting, and their mRNA levels were examined using quantitative real-time reverse transcription-polymerase chain reaction. RESULTS: WHF treatment was found to significantly ameliorate weight loss, proteinuria, hypoalbuminemia, and dyslipidemia in doxorubicin-nephropathy rats. The protein and mRNA levels of TRPC6, STIM1, and Orai1 were partially, but significantly suppressed by prednisone or WHF treatment. CONCLUSION: Treatment with WHF significantly ameliorates renal injury in a rat model of doxorubicin-induced nephropathy, which could be at least partially related to repression of the TRPC6 pathway.
Subject(s)
Doxorubicin/adverse effects , Drugs, Chinese Herbal/administration & dosage , Kidney Diseases/prevention & control , Protective Agents/administration & dosage , TRPC Cation Channels/metabolism , Animals , Disease Models, Animal , Humans , Kidney/drug effects , Kidney/metabolism , Kidney Diseases/chemically induced , Kidney Diseases/genetics , Kidney Diseases/metabolism , Male , Rats , Rats, Sprague-Dawley , TRPC Cation Channels/geneticsABSTRACT
Chrysin (CH) is the main ingredient of many medicinal plants. Our previous study showed that CH could suppress hypoxia-induced pulmonary arterial smooth muscle cells proliferation and alleviate chronic hypoxia-induced pulmonary hypertension by targeting store-operated Ca entry (SOCE)-[Ca]i pathway. In this study, we investigated the effect of CH on monocrotaline-induced pulmonary hypertension (MCTPH) and the mechanism behind it. Results show that, in MCTPH model rats, (1) CH significantly reduced the enhancement of right ventricular pressure, right ventricular hypertrophy, and pulmonary vascular remodeling; (2) CH markedly suppressed the promotion of SOCE and [Ca]i in pulmonary arterial smooth muscle cells; and (3) CH obviously inhibited the MCT-upregulated proliferating cell nuclear antigen, TRPC1, TRPC4, and TRPC6 expression in distal pulmonary arteries. These results demonstrate that CH likely alleviates MCTPH by targeting TRPC1,4,6-SOCE-[Ca]i pathway.
Subject(s)
Antihypertensive Agents/pharmacology , Arterial Pressure/drug effects , Calcium Signaling/drug effects , Flavonoids/pharmacology , Hypertension, Pulmonary/drug therapy , Monocrotaline , Muscle, Smooth, Vascular/drug effects , Vascular Remodeling/drug effects , Animals , Disease Models, Animal , Hypertension, Pulmonary/chemically induced , Hypertension, Pulmonary/metabolism , Hypertension, Pulmonary/physiopathology , Hypertrophy, Right Ventricular/chemically induced , Hypertrophy, Right Ventricular/metabolism , Hypertrophy, Right Ventricular/physiopathology , Hypertrophy, Right Ventricular/prevention & control , Muscle, Smooth, Vascular/metabolism , Muscle, Smooth, Vascular/physiopathology , Pulmonary Artery/drug effects , Pulmonary Artery/metabolism , Pulmonary Artery/physiopathology , Rats, Sprague-Dawley , TRPC Cation Channels/antagonists & inhibitors , TRPC Cation Channels/metabolism , Ventricular Function, Right/drug effects , Ventricular Pressure/drug effects , Ventricular Remodeling/drug effectsABSTRACT
Cortex Mori Radicis extract (CMR) has various pharmacological properties, such as antiinflammatory, antiallergic and antihyperglycemic effects. However, the effects and mechanisms of CMR in the neuroregeneration of diabetic peripheral neuropathy (DPN) are unclear. In the present study, the effects of CMR on neurite outgrowth of dorsal root ganglia (DRG) neurons in diabetic rats were investigated and its underlying mechanisms were explored. SD rats were subjected to a highfat diet with lowdose streptozotocin to induce a Type II diabetes model with peripheral neuropathy. CMR was then applied for four weeks continuously with or without injection of small interfere (si)RNA targeting the transient receptor potential canonical channel 1 (TRPC1) via the tail vein. Blood glucose levels, the number of Nissl bodies, neurite outgrowth and growth cone turning in DRG neurons were evaluated. The expression of TRPC1 protein, Ca2+ influx and activation of the PI3K/AKT signaling pathway were also investigated. The results of the present study showed that CMR significantly lowered blood glucose levels, reversed the loss of Nissl bodies, induced neurite outgrowth and restored the response of the growth cone of DRG neurons in diabetic rats. CMR exerted neurite outgrowthpromoting effects by increasing TRPC1 expression, reducing Ca2+ influx and enhancing AKT phosphorylation. siRNA targeting TRPC1 in the CMR group abrogated its antidiabetic and neuroregenerative effects, suggesting the involvement of TRPC1 in the biological effects of CMR on DPN.
Subject(s)
Diabetes Mellitus, Experimental/drug therapy , Diabetic Neuropathies/drug therapy , Morus , Neurites/metabolism , Neuronal Outgrowth/drug effects , Plant Extracts/pharmacology , Animals , Blood Glucose/drug effects , Calcium/metabolism , Cells, Cultured , Diabetes Mellitus, Experimental/blood , Diabetes Mellitus, Experimental/genetics , Diabetes Mellitus, Experimental/metabolism , Diabetic Neuropathies/blood , Diabetic Neuropathies/genetics , Diabetic Neuropathies/metabolism , Drugs, Chinese Herbal/pharmacology , Drugs, Chinese Herbal/therapeutic use , Ganglia, Spinal/drug effects , Ganglia, Spinal/growth & development , Ganglia, Spinal/metabolism , Male , Neurites/drug effects , Neurites/pathology , Neurons/drug effects , Neurons/metabolism , Nissl Bodies/drug effects , Nissl Bodies/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Phosphorylation , Plant Extracts/therapeutic use , Proto-Oncogene Proteins c-akt/metabolism , RNA, Small Interfering , Rats , Rats, Sprague-Dawley , Signal Transduction/drug effects , Signal Transduction/genetics , TRPC Cation Channels/genetics , TRPC Cation Channels/metabolism , Up-RegulationABSTRACT
Chrysin (CH), the main ingredient of many medicinal plants, has been reported to be a very potent flavonoid possessing a large number of pharmacological activities. Recent studies have shown that CH significantly improves hemodynamic parameters such as right ventricular pressure, right ventricular hypertrophy, and pulmonary vascular remodeling in a rat model of chronic hypoxia-induced pulmonary hypertension (CHPH). These improvements are through the inhibition of NOX4 expression, reactive oxygen species and malondialdehyde production, pulmonary arterial smooth muscle cell (PASMC) proliferation, and collagen accumulation. In this study, we investigated another mechanism by which CH alleviates CHPH by regulating intracellular calcium concentrations ([Ca]i) in PASMCs, as well as the underlying signaling pathway. The results show that (1) in CHPH model rats, CH substantially attenuated elevated right ventricular pressure, right ventricular hypertrophy, and pulmonary vascular remodeling; (2) in cultured rat distal PASMCs, CH inhibited the hypoxia-triggered promotion of cell proliferation, store-operated Ca entry and [Ca]i; and (3) CH significantly suppressed the hypoxia-upregulated HIF-1α, BMP4, TRPC1, and TRPC6 expression in distal pulmonary arteries (PAs) and cultured rat distal PASMCs. These results indicate that CH likely exerts its CHPH protective activity by regulating [Ca]i, which may result from the downregulation of HIF-1α, BMP4, TRPC1, and TRPC in PASMCs.
Subject(s)
Antihypertensive Agents/pharmacology , Calcium Signaling/drug effects , Calcium/metabolism , Flavonoids/pharmacology , Hypertension, Pulmonary/prevention & control , Hypoxia/drug therapy , Muscle, Smooth, Vascular/drug effects , Myocytes, Smooth Muscle/drug effects , Animals , Bone Morphogenetic Protein 4/metabolism , Cell Proliferation/drug effects , Cells, Cultured , Hypertension, Pulmonary/etiology , Hypertension, Pulmonary/metabolism , Hypertension, Pulmonary/physiopathology , Hypoxia/complications , Hypoxia/metabolism , Hypoxia/physiopathology , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Muscle, Smooth, Vascular/metabolism , Muscle, Smooth, Vascular/physiopathology , Myocytes, Smooth Muscle/metabolism , Pulmonary Artery/drug effects , Pulmonary Artery/metabolism , Pulmonary Artery/physiopathology , Rats, Sprague-Dawley , TRPC Cation Channels/metabolism , Vascular Remodeling/drug effects , Ventricular Function, Right/drug effects , Ventricular Pressure/drug effects , Ventricular Remodeling/drug effectsABSTRACT
We show that both supplemental and ambient magnetic fields modulate myogenesis. A lone 10 min exposure of myoblasts to 1.5 mT amplitude supplemental pulsed magnetic fields (PEMFs) accentuated in vitro myogenesis by stimulating transient receptor potential (TRP)-C1-mediated calcium entry and downstream nuclear factor of activated T cells (NFAT)-transcriptional and P300/CBP-associated factor (PCAF)-epigenetic cascades, whereas depriving myoblasts of ambient magnetic fields slowed myogenesis, reduced TRPC1 expression, and silenced NFAT-transcriptional and PCAF-epigenetic cascades. The expression levels of peroxisome proliferator-activated receptor γ coactivator 1α, the master regulator of mitochondriogenesis, was also enhanced by brief PEMF exposure. Accordingly, mitochondriogenesis and respiratory capacity were both enhanced with PEMF exposure, paralleling TRPC1 expression and pharmacological sensitivity. Clustered regularly interspaced short palindromic repeats-Cas9 knockdown of TRPC1 precluded proliferative and mitochondrial responses to supplemental PEMFs, whereas small interfering RNA gene silencing of TRPM7 did not, coinciding with data that magnetoreception did not coincide with the expression or function of other TRP channels. The aminoglycoside antibiotics antagonized and down-regulated TRPC1 expression and, when applied concomitantly with PEMF exposure, attenuated PEMF-stimulated calcium entry, mitochondrial respiration, proliferation, differentiation, and epigenetic directive in myoblasts, elucidating why the developmental potential of magnetic fields may have previously escaped detection. Mitochondrial-based survival adaptations were also activated upon PEMF stimulation. Magnetism thus deploys an authentic myogenic directive that relies on an interplay between mitochondria and TRPC1 to reach fruition.-Yap, J. L. Y., Tai, Y. K., Fröhlich, J., Fong, C. H. H., Yin, J. N., Foo, Z. L., Ramanan, S., Beyer, C., Toh, S. J., Casarosa, M., Bharathy, N., Kala, M. P., Egli, M., Taneja, R., Lee, C. N., Franco-Obregón, A. Ambient and supplemental magnetic fields promote myogenesis via a TRPC1-mitochondrial axis: evidence of a magnetic mitohormetic mechanism.
Subject(s)
Magnetic Fields , Mitochondria, Muscle/metabolism , Muscle Development , Myoblasts, Skeletal/metabolism , Signal Transduction , TRPC Cation Channels/metabolism , Animals , Cell Line , Mice , Mitochondria, Muscle/genetics , Myoblasts, Skeletal/cytology , TRPC Cation Channels/geneticsABSTRACT
Enhancing soluble receptor for advanced glycation endproducts (sRAGE) is considered as a potent strategy for diabetes therapy. sRAGE secretion is regulated by calcium and transient receptor potential canonical (TRPC) channels. However, the role of TRPC channels in diabetes remains unknown. 18ß-Glycyrrhetinic acid (18ß-GA), produced from liquorice, has shown antidiabetic properties. This study was aimed to investigate the effect of 18ß-GA on sRAGE secretion via TRPC channels in high glucose (HG)-induced THP-1 cells. HG treatment enhanced TRPC3 and TRPC6 expression and consequently caused reactive oxygen species (ROS) accumulation mediated through p47 nicotinamide-adenine dinucleotide phosphate oxidase and inducible nitric oxide synthase (iNOS) associated with uncoupling protein 2 (UCP2) decline and lower sRAGE secretion. Interestingly, 18ß-GA showed the dramatic effects similar to Pyr3 or 2-aminoethyl diphenyl borinate inhibitors and effectively reversed HG-elicited mechanisms including that blocking TRPC3 and TRPC6 protein expressions, suppressing intracellular [Ca2+] concentration, decreasing expressions of ROS, p47s, and iNOS, but increasing UCP2 level and promoting sRAGE secretion. Therefore, 18ß-GA provides a potential implication to diabetes mellitus and its complications.
Subject(s)
Glucose/antagonists & inhibitors , Glycyrrhetinic Acid/analogs & derivatives , Glycyrrhiza/chemistry , Hypoglycemic Agents/pharmacology , Receptor for Advanced Glycation End Products/genetics , TRPC Cation Channels/genetics , TRPC6 Cation Channel/genetics , Boron Compounds/pharmacology , Calcium/metabolism , Gene Expression Regulation , Glucose/toxicity , Glycyrrhetinic Acid/isolation & purification , Glycyrrhetinic Acid/pharmacology , Humans , Hypoglycemic Agents/isolation & purification , NADPH Oxidases/genetics , NADPH Oxidases/metabolism , Nitric Oxide Synthase Type II/genetics , Nitric Oxide Synthase Type II/metabolism , Plant Extracts/chemistry , Pyrazoles/pharmacology , Reactive Oxygen Species/antagonists & inhibitors , Reactive Oxygen Species/metabolism , Receptor for Advanced Glycation End Products/antagonists & inhibitors , Receptor for Advanced Glycation End Products/metabolism , Signal Transduction , THP-1 Cells , TRPC Cation Channels/antagonists & inhibitors , TRPC Cation Channels/metabolism , TRPC6 Cation Channel/antagonists & inhibitors , TRPC6 Cation Channel/metabolism , Uncoupling Protein 2/genetics , Uncoupling Protein 2/metabolismABSTRACT
Alzheimer's disease (AD) is a neurodegenerative disorder that is the major cause of dementia in the elderly. There is no cure against AD. We have recently discovered a novel transient receptor potential canonical 6 (TRPC6)-mediated intracellular signaling pathway that regulates the stability of dendritic spines and plays a role in memory formation. We have previously shown that TRPC6 agonists exert beneficial effects in models of AD and may serve as lead compounds for development of AD therapeutic agents. In the current study, we used the Clarivate Analytics Integrity database to search for additional TRPC6 agonists. We selected four compounds to study as potential neuroprotective agents. We applied bioinformatics analyses to test the basic pharmacological properties of the selected compounds. We performed in vitro screening of these compounds to validate their ability to protect mushroom spines from amyloid toxicity and determined that two of these compounds exert neuroprotective effects in the nanomolar concentration range. We have chosen one of these compounds [piperazine (PPZ)] for further testing. In agreement with previously published data, we have shown that PPZ potentiates TRPC6 channels. We demonstrated that the neuroprotective mechanism of the investigated PPZ is based on activation of neuronal store-operated calcium entry in spines. We have shown that PPZ restores long-term potentiation induction in 6-month-old 5xFAD mouse hippocampal slices. The obtained results suggest that PPZ and its derivatives are potential lead molecules for development of AD therapeutic agents.
Subject(s)
Alzheimer Disease/drug therapy , Piperazines/pharmacology , Alzheimer Disease/metabolism , Animals , Calcium Signaling/drug effects , Dendritic Spines/drug effects , Dendritic Spines/metabolism , Disease Models, Animal , Hippocampus/drug effects , Hippocampus/metabolism , Long-Term Potentiation/drug effects , Mice , Mice, Inbred C57BL , Neurons/drug effects , Neurons/metabolism , Neuroprotective Agents/pharmacology , TRPC Cation Channels/metabolismABSTRACT
Reactive oxygen species induce vascular dysfunction and hypertension by directly interacting with nitric oxide (NO) which leads to NO inactivation. In addition to a decrease in NO bioavailability, there is evidence that oxidative stress can also modulate NO signaling during hypertension. Here, we investigated the effect of oxidative stress on NO signaling molecules cGMP-dependent protein kinase (PKG) and vasodilator-stimulated phosphoprotein (VASP) which are known to mediate vasodilatory actions of NO. Male Sprague Dawley (SD) rats were provided with tap water (control), 30 mM L-buthionine sulfoximine (BSO, a pro-oxidant), 1 mM tempol (T, an antioxidant) and BSO + T for 3 wks. BSO-treated rats exhibited high blood pressure and oxidative stress. Incubation of mesenteric arterial rings with NO donors caused concentration-dependent relaxation in control rats. However, the response to NO donors was significantly lower in BSO-treated rats with a marked decrease in pD2. In control rats, NO donors activated mesenteric PKG, increased VASP phosphorylation and its interaction with transient receptor potential channels 4 (TRPC4) and inhibited store-operated Ca2+ influx. NO failed to activate these signaling molecules in mesenteric arteries from BSO-treated rats. Supplementation of BSO-treated rats with tempol reduced oxidative stress and blood pressure and normalized the NO signaling. These data suggest that oxidative stress can reduce NO-mediated PKG activation and VASP-TRPC4 interaction which leads to failure of NO to reduce Ca2+ influx in smooth muscle cells. The increase in intracellular Ca2+ contributes to sustained vasoconstriction and subsequent hypertension. Antioxidant supplementation decreases oxidative stress, normalizes NO signaling and reduces blood pressure.
Subject(s)
Cell Adhesion Molecules/metabolism , Cyclic GMP-Dependent Protein Kinases/metabolism , Hypertension/physiopathology , Mesenteric Arteries/physiopathology , Microfilament Proteins/metabolism , Nitric Oxide/metabolism , Oxidative Stress , Phosphoproteins/metabolism , Animals , Antioxidants/pharmacology , Blood Pressure/drug effects , Buthionine Sulfoximine/pharmacology , Calcium/metabolism , Cyclic N-Oxides/pharmacology , Male , Mesenteric Arteries/drug effects , Oxidative Stress/drug effects , Phosphorylation , Rats , Rats, Sprague-Dawley , Reactive Oxygen Species/pharmacology , Serine/metabolism , Signal Transduction/drug effects , Spin Labels , TRPC Cation Channels/metabolism , Vasodilation/drug effectsABSTRACT
Iodine is an essential component for thyroid hormone synthesis. Epidemiological investigations have demonstrated that maternal mild iodine deficiency (ID)-induced hypothyroxinemia can affect intellectual and behavioral function in offspring. There is no definitive evidence demonstrating the effects of maternal iodine supplementation on neurobehavioral function in regional areas with mild ID. Thus, we aimed to clarify the effects of maternal mild ID and iodine supplementation on motor coordination in offspring and illuminate the underlying molecular mechanisms. Animal models of maternal mild ID and iodine supplementation were generated by providing Wistar rats an iodine-deficient diet and deionized water supplemented with potassium iodide during pregnancy and lactation. We found that mild ID-induced hypothyroxinemia led to a shorter latent time before falling down from the rotarod, a longer time to traverse the balance beam and poorer wire grip of the forelimbs, which imply motor coordination dysfunction. However, these impairments in the offspring were improved by iodine supplementation during pregnancy and lactation. We further observed that the ultrastructure and dendritic tree morphology of cerebellar Purkinje cells were altered in mild ID-induced hypothyroxinemia but that these changes could be reversed by iodine supplementation. Maternal mild ID and iodine supplementation also affected expression of the mGluR1 signaling pathway in offspring. Together, iodine supplementation during pregnancy and lactation can improve motor coordination in offspring by modulating the mGluR1 signaling pathway in mild ID-induced hypothyroxinemia rats.
Subject(s)
Iodine/pharmacology , Motor Activity/drug effects , Receptors, Metabotropic Glutamate/metabolism , Thyroxine/blood , Animals , Dietary Supplements , Female , Iodine/deficiency , Iodine/urine , Lactation , Male , Maternal Nutritional Physiological Phenomena , Phospholipase D/metabolism , Pregnancy , Purkinje Cells/drug effects , Rats, Wistar , Rotarod Performance Test , Signal Transduction/drug effects , TRPC Cation Channels/metabolism , Thyroid Hormones/bloodABSTRACT
All of the canonical transient receptor potential channels (TRPC) with the exception of TRPC 2 are expressed in hypothalamic neurons and are involved in multiple homeostatic functions. Although the metabotropic glutamate receptors have been shown to be coupled to TRPC channel activation in cortical and sub-cortical brain regions, in the hypothalamus multiple amine and peptidergic G protein-coupled receptors (GPCRs) and growth factor/cytokine receptors are linked to activation of TRPC channels that are vital for reproduction, temperature regulation, arousal and energy homeostasis. In addition to the neurotransmitters, circulating hormones like insulin and leptin through their cognate receptors activate TRPC channels in POMC neurons. Many of the post-synaptic effects of the neurotransmitters and hormones are regulated in different physiological states by expression of TRPC channels in the post-synaptic neurons. Therefore, TRPC channels are key targets not only for neurotransmitters but circulating hormones in their vital role to control multiple hypothalamic functions, which is the focus of this review.
Subject(s)
Gonadotropin-Releasing Hormone/metabolism , Hypothalamus/metabolism , Insulin/metabolism , Intercellular Signaling Peptides and Proteins/metabolism , Neurons/metabolism , Orexins/metabolism , Pro-Opiomelanocortin/metabolism , Receptors, G-Protein-Coupled/metabolism , TRPC Cation Channels/metabolism , Animals , HumansABSTRACT
Because of the serious side effects of the currently used bronchodilators, new compounds with similar functions must be developed. We screened several herbs and found that Polygonum aviculare L. contains ingredients that inhibit the precontraction of mouse and human airway smooth muscle (ASM). High K+-induced precontraction in ASM was completely inhibited by nifedipine, a selective blocker of L-type voltage-dependent Ca2+ channels (LVDCCs). However, nifedipine only partially reduced the precontraction induced by acetylcholine chloride (ACH). Additionally, the ACH-induced precontraction was partly reduced by pyrazole-3 (Pyr3), a selective blocker of TRPC3 and stromal interaction molecule (STIM)/Orai channels. These channel-mediated currents were inhibited by the compounds present in P. aviculare extracts, suggesting that this inhibition was mediated by LVDCCs, TRPC3 and/or STIM/Orai channels. Moreover, these channel-mediated currents were inhibited by quercetin, which is present in P. aviculare extracts. Furthermore, quercetin inhibited ACH-induced precontraction in ASM. Overall, our data indicate that the ethyl acetate fraction of P. aviculare and quercetin can inhibit Ca2+-permeant LVDCCs, TRPC3 and STIM/Orai channels, which inhibits the precontraction of ASM. These findings suggest that P. aviculare could be used to develop new bronchodilators to treat obstructive lung diseases such as asthma and chronic obstructive pulmonary disease.
Subject(s)
Muscle Contraction/drug effects , Muscle, Smooth/drug effects , Plant Extracts/pharmacology , Polygonum/chemistry , Quercetin/pharmacology , Acetylcholine/pharmacology , Animals , Calcium/metabolism , Calcium Channels, L-Type/metabolism , Humans , Lung/drug effects , Lung/metabolism , Male , Mice , Mice, Inbred BALB C , Muscle, Smooth/metabolism , Nifedipine/pharmacology , TRPC Cation Channels/metabolismABSTRACT
To investigate the inhibitory effect of Huangqi Danshen decoction (HDD) on isoproterenol (ISO)-induced myocardial remodeling and explore its effect on STIM1, TRPC1, CaN and NFATc3 expressions. ISO (2.5 mgâ¢kg⻹â¢d⻹×14 d) was given by subcutaneous injection to establish myocardial remodeling models in rats, and then were randomly divided into control group, ISO model group, HDD5 group (HDD 5 gâ¢kg⻹â¢d⻹+ISO), and HDD10 group (HDD 10 gâ¢kg⻹â¢d⻹+ISO). After intervention for 4 weeks, the heart mass index (HW/BW) and the left ventricular mass index (LVW/BW) were calculated; the structure of myocardium was observed by echocardiography; the pathological changes of myocardium were observed by HE staining; levels of BNP, CaN and CaM kinases II in serum were detected by ELISA, and the protein expression levels of STIM1, TRPC1, p-CaN, p-NFATc3, and NFATc3 in left ventricular tissues were detected by Western blot. The results showed that the HW/BW and LVW/BW in ISO group were greater than those in HDD5 group and HDD10 group (P<0.05); Echocardiography showed that HDD inhibited ISO-induced increase in LVEDD and LVESD; ELISA results showed that HDD could significantly inhibit the increase of BNP, CaN and CaM kinases II levels in serum of rats with ISO-induced myocardial remodeling (P<0.01). Western blot results showed that STIM1, TRPC1, p-CaN, p-NFATc3 and NFATc3 expression levels were increased in the myocardial tissues of ISO group rats, and after HDD administration, the above expression levels were decreased in group ISO, HDD for myocardial tissue after administration of STIM1, TRPC1, p-CaN, p-NFATc3 and NFATc3 expression decreased (P<0.05). Our findings indicated that HDD can attenuate the myocardial remodeling induced by ISO, and its mechanism may be related to down-regulating the expression levels of STIM1, TRPC1, CaM kinases II, p-CaN/CaN and p-NFATc3/NFATc3.
Subject(s)
Drugs, Chinese Herbal/pharmacology , Heart/drug effects , NFATC Transcription Factors/metabolism , Salvia miltiorrhiza/chemistry , Stromal Interaction Molecule 1/metabolism , TRPC Cation Channels/metabolism , Ventricular Remodeling , Animals , Calcium-Calmodulin-Dependent Protein Kinase Type 2/metabolism , Isoproterenol , Myocardium , Random Allocation , Rats , Rats, Sprague-Dawley , Rats, WistarABSTRACT
CONTEXT: Fructus Psoralea, Psoralea corylifolia L. (Leguminosae), has been widely used in traditional medicines for the treatment of dermatitis, leukoderma, asthma and osteoporosis. OBJECTIVES: In this study, we sought to study mechanisms underlying the vasoactive properties of Psoralea corylifolia extract (PCE) and its active ingredients. MATERIALS AND METHODS: To study mechanisms underlying the vasoactive properties of PCE prepared by extracting dried seeds of Psoralea corylifolia with 70% ethanol, isometric tension recordings of rat aortic rings and the ionic currents through TRPC3 (transient receptor potential canonical 3) channels were measured with the cumulative concentration (10-600 µg/mL) of PCE or its constituents. RESULTS: Cumulative treatment with PCE caused the relaxation of pre-contracted aortic rings in the presence and absence of endothelium with EC50 values of 61.27 ± 3.11 and 211.13 ± 18.74 µg/mL, respectively. Pretreatment with inhibitors of nitric oxide (NO) synthase, guanylate cyclase, or cyclooxygenase and pyrazole 3, a selective TRPC3 channel blocker, significantly decreased PCE-induced vasorelaxation (p < 0.01). The PCE constituents, bakuchiol, isobavachalcone, isopsoralen and psoralen, inhibited hTRPC3 currents (inhibited by 40.6 ± 2.7, 27.1 ± 7.9, 35.1 ± 4.8 and 47.4 ± 3.9%, respectively). Furthermore, these constituents significantly relaxed pre-contracted aortic rings (EC50 128.9, 4.5, 32.1 and 114.9 µg/mL, respectively). DISCUSSION AND CONCLUSIONS: Taken together, our data indicate that the vasodilatory actions of PCE are dependent on endothelial NO/cGMP and also involved in prostaglandin production. PCE and its active constituents, bakuchiol, isobavachalcone, isopsoralen and psoralen, caused dose-dependent inhibition of TRPC3 channels, indicating that those ingredients attenuate Phe-induced vasoconstriction.