[Rapid health technology assessment of four oral Chinese patent medicines in treatment of functional gastrointestinal disorders].

To provide proof of the evidence-based medicine and decision-making information for the clinical decision of functional gastrointestinal disorders(FGIDs), this study evaluated and compared the efficacy, safety, and economy of four oral Chinese patent medicines(CPMs) in the treatment of FGIDs using the method of rapid health technology assessment. The literature was systematically retrieved from CNKI, Wanfang, VIP, SinoMed, EMbase, PubMed, Cochrane Library and ClinicalTrials.gov from the establishment of the databases to May 1, 2022. Two evaluators screened out the literature, extracted data, evaluated the quality of the literature, and descriptively analyzed the results according to the prepared standard. Eventually, 16 studies were included, all of which was rando-mized controlled trial(RCT). The results showed that Renshen Jianpi Tablets, Renshen Jianpi Pills, Shenling Baizhu Granules, and Buzhong Yiqi Granules all had certain effects on the treatment of FGIDs. Renshen Jianpi Tablets treated FGIDs and persistent diarrhea. Shenling Baizhu Granules treated diarrhea with irritable bowel syndrome and FGIDs. Buzhong Yiqi Granules treated diarrhea with irritable bowel syndrome, FGIDs, and chronic diarrhea in children. Renshen Jianpi Pills treated chronic diarrhea. The four oral CPMs all have certain effects on the treatment of FGIDs and have specific advantages for specific patients. Compared with other CPMs, Renshen Jianpi Tablets have higher clinical universality. However, there are problems such as insufficient clinical research evidence, generally low quality of evidence, lack of comparative analysis among medicines, and lack of academic evaluation. More high-quality clinical research and the economic research should be carried out in the future, so as to provide more evidence for the evaluation of the four CPMs.

Wheat Bax Inhibitor-1 interacts with TaFKBP62 and mediates response to heat stress.

BACKGROUND: Heat stress is a severe environmental stress that affects plant growth and reduces yield. Bax inhibitor-1 (BI-1) is a cytoprotective protein that is involved in the response to biotic and abiotic stresses. The Arabidopsis (Arabidopsis thaliana) BI-1 mutants atbi1-1 and atbi1-2 are hypersensitive to heat stress, and AtBI-1 overexpression rescues thermotolerance deficiency in atbi1 plants. Nevertheless, the mechanism of BI-1 in plant thermotolerance is still unclear. RESULTS: We identified a wheat (Triticum aestivum L.) BI-1 gene, TaBI-1.1, which was highly upregulated in an RNA sequencing (RNA-seq) analysis of heat-treated wheat. The upregulation of TaBI-1.1 under heat stress was further demonstrated by real time quantitative PCR (qRT-PCR) and ß-glucuronidase (GUS) staining. Compared with the wild type Col-0, the atbi1-2 mutant is hypersensitive to heat stress, and constitutive expression of TaBI-1.1 in atbi1-2 (35S::TaBI-1.1/ atbi1-2) rescued the deficiency of atbi1-2 under heat stress. Furthermore, we identified TaFKBP62 as a TaBI-1.1-interacting protein that co-localized with TaBI-1.1 on the endoplasmic reticulum (ER) membrane and enhanced heat stress tolerance. Additionally, HSFA2, HSFB1, ROF1, HSP17.4B, HSP17.6A, HSP17.8, HSP70B, and HSP90.1 expression levels were suppressed in atbi1-2 plants under heat stress. In contrast, 35S::TaBI-1.1/atbi1-2 relieved the inhibitory effect of AtBI-1 loss of function. CONCLUSIONS: TaBI-1.1 interacted with TaFKBP62 and co-localized with TaFKBP62 on the ER membrane. Both TaBI-1.1 and AtBI-1 regulated the expression of heat-responsive genes and were conserved in plant thermotolerance.

The Protective Effect of Propofol Against Ischemia-Reperfusion Injury in the Interlobar Arteries: Reduction of Abnormal Cx43 Expression as a Possible Mechanism.

BACKGROUND/AIMS: This experimental study aims to observe whether the protective effect of propofol against renal ischemia-reperfusion injury (IRI) in the rat interlobar artery occurs through altered expression of the gap junction protein connexin 43 (Cx43). METHODS: This study randomly divided male Sprague Dawley (SD) rats into an untreated control group, a sham-operated control group (sham group), an ischemia-reperfusion group (IR group), a propofol group (propofol+IR group) and a fat emulsion group (Intralipid group). The ischemia/reperfusion model was prepared through resection of the right kidney and noninvasive arterial occlusion of the left kidney. Forty-five minutes after renal ischemia-reperfusion, an automatic biochemical analyzer was employed to measure blood urea nitrogen (BUN) and serum creatinine (SCr); changes in renal tissue pathology were observed using hematoxylin and eosin (HE) staining, and the vasomotor activity of the interlobar artery was detected using a pressure mechanogram technique. The protein expression of Cx43 in renal artery cross-sections was determined through western blotting. RESULTS: The experimental study confirmed that the BUN and SCr of rats markedly increased after ischemia-reperfusion injury; additionally, we observed some coagulation necrosis and shedding of cells, some solidification of nuclear chromatin, degeneration of cytoplasmic vacuoles, high renal interstitial vascular congestion and obvious inflammatory cell infiltration, characterized by focal hemorrhages. Furthermore, the contraction activity of the renal interlobar artery greatly decreased, and the tension of the arteries in the renal lobe increased remarkably. After the gap junction blocking agents 2-APB and Gap27 were applied, the systolic velocity of blood vessels and the vascular contraction rate both decreased. In addition, the expression of Cx43 in kidney tissues increased markedly. The damage was more severe after 24 h of ischemic reperfusion than after only 4 h. However, after pretreatment with propofol, regardless of whether ischemia-reperfusion was applied for 4 h or 24 h, the previously increased expression of Cx43 decreased obviously, and all forms of renal damage were reversed. CONCLUSION: Our research suggests new ways for propofol to relieve ischemia-reperfusion injury by decreasing the abnormal expression of the gap junction protein Cx43. This study reveals a novel mechanism for the action of propofol against IRI, and we hope this finding will lead to new treatments for IRI.

The WRKY Transcription Factor GmWRKY12 Confers Drought and Salt Tolerance in Soybean.

WRKYs are important regulators in plant development and stress responses. However, knowledge of this superfamily in soybean is limited. In this study, we characterized the drought- and salt-induced gene GmWRKY12 based on RNA-Seq and qRT-PCR. GmWRKY12, which is 714 bp in length, encoded 237 amino acids and grouped into WRKY II. The promoter region of GmWRKY12 included ABER4, MYB, MYC, GT-1, W-box and DPBF cis-elements, which possibly participate in abscisic acid (ABA), drought and salt stress responses. GmWRKY12 was minimally expressed in different tissues under normal conditions but highly expressed under drought and salt treatments. As a nucleus protein, GmWRKY12 was responsive to drought, salt, ABA and salicylic acid (SA) stresses. Using a transgenic hairy root assay, we further characterized the roles of GmWRKY12 in abiotic stress tolerance. Compared with control (Williams 82), overexpression of GmWRKY12 enhanced drought and salt tolerance, increased proline (Pro) content and decreased malondialdehyde (MDA) content under drought and salt treatment in transgenic soybean seedlings. These results may provide a basis to understand the functions of GmWRKY12 in abiotic stress responses in soybean.

Enhanced gap junctional channel activity between vascular smooth muscle cells in cerebral artery of spontaneously hypertensive rats.

The aim of the present study is to investigate the effects of hypertension on the gap junctions between vascular smooth muscle cells (VSMCs) in the cerebral arteries (CAs) of spontaneously hypertensive rats (SHRs). The functions of gap junctions in the CAs of VSMCs in SHRs and control normotensive Wistar-Kyoto (WKY) rats were studied using whole-cell patch clamp recordings and pressure myography, and the expression levels of connexins were analyzed using reverse transcription-quantitative polymerase chain reaction and Western blot analyses. Whole-cell patch clamp measurements revealed that the membrane capacitance and conductance of in situ VSMCs in the CAs were significantly greater in SHRs than in WKY rats, suggesting that gap junction coupling is enhanced between VSMCs in the CAs of SHRs. Application of the endothelium-independent vasoconstrictors KCl or phenylephrine (PE) stimulated a greater vasoconstriction in the CAs of SHRs than in those of WKY rats. The EC50 value of KCl was 24.9 mM (n = 14) and 36.9 mM (n=12) for SHRs and WKY rats, respectively. The EC50 value of PE was 0.9 µM (n = 7) and 2.2 µM (n = 7) for SHRs and WKY rats, respectively. Gap junction inhibitors 18ß-glycyrrhetinic acid (18ß-GA), niflumic acid (NFA), and 2-aminoethoxydiphenyl borate (2-APB) attenuated KCl-induced vasoconstriction in SHRs and WKY rats. The mRNA and protein expression levels of the gap junction protein connexin 45 (Cx45) were significantly higher in the CAs of SHRs than in those of WKY rats. Phosphorylated Cx43 protein expression was significantly higher in the CAs of SHRs than in those of WKY rats, despite the total Cx43 mRNA and protein expression levels in the cerebral artery (CA) exhibiting no significant difference between SHRs and WKY rats. Increases in the expression of Cx45 and phosphorylation of Cx43 may promote gap junction communication among VSMCs in the CAs of SHRs, which may enhance the contractile response of the CA to vasoconstrictors.

The enhancement of Cx45 expression and function in renal interlobar artery of spontaneously hypertensive rats at different age.

BACKGROUND/AIMS: This study was designed to investigate the expression and function of gap junction protein connexin 45 (Cx45) in renal interlobar artery (RIA) of spontaneously hypertensive rats (SHR), and the association between hypertension and enhanced vasoconstrictive response in SHR. METHODS: Western blot analysis and pressure myography were used to examine the differences in expression and function of Cx45 in vascular smooth muscle cells (VSMCs) of RIA between SHR and normotensive Wistar-Kyoto (WKY) rats. RESULTS: Our results demonstrated that 1) whole-cell patch clamp measurements showed that the membrane capacitance and conductance of in-situ RIA VSMCs of SHR were significantly greater than those of WKY rats (p<0.05, n=6), suggesting that the coupling of gap junction between VSMCs of RIA was enhanced in SHR; 2) the KCl or phenylephrine (PE)-stimulated RIA constriction was more pronounced in SHR than that in WKY rats (p<0.05, n=10). After applying a gap junction inhibitor 18ß-glycyrrhetintic acid (18ß-GA), the inhibitory effect of 18ß-GA on KCl or PE-induced vasoconstriction was greater in SHR (p<0.05, n=10); and 3) the expression of Cx45 in RIA of SHR was greater than that in WKY rats (p<0.05, n=3) at 4, 12 and 48 wks of age. CONCLUSIONS: The hypertension-induced elevation of Cx45 may affect communication between VSMCs and coupling between VSMCs and endothelium, which results in an increased vasoconstrictive response in renal artery and might contribute to the development of hypertension.

PKCɛ mediates substance P inhibition of GABAA receptors-mediated current in rat dorsal root ganglion.

The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1-1000 µmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1-1000 µmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001-1 µmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 µmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCß. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca²âº-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the "pre-synaptic inhibition" evoked by GABA, which may explain its role in pain and neurogenic inflammation.

Myoendothelial coupling is unidirectional in guinea pig spiral modiolar arteries.

Gap junctions (GJs) facilitate communication and promote transfer of signaling molecules or current between adjacent cells in various organs to coordinate cellular activity. In arteries, homocellular GJs are present between adjacent smooth muscle cells (SMCs) and between adjacent endothelial cells (ECs), whilst many arteries also exhibit heterocellular GJs between SMCs and ECs. To test the hypothesis that there is differential cell coupling in guinea pig spiral modiolar arteries (SMA), we used intracellular recording technique to record cellular activities simultaneously in ECs or SMCs in acutely isolated guinea pig SMA preparations. Cell types were identified by injection of a fluorescent dye, propidium iodide (PI), through recording microelectrodes. Stable intracellular recordings were made in 120 cells among which 61 were identified as SMCs and 28 as ECs. Dual intracellular recordings were conducted to detect the coexistence of the two distinct levels of resting potential (RP) and to estimate the intensity of electrical coupling between two cells by a current pulse of up to 0.5-1.5 nA. The electrotonic potential was detected not only in the current-injected cell, but also in the majority of non-injected cells. The electrical coupling ratios (ECRs) of homocellular cells were not significant (P>0.05) (0.084±0.032 (n=6) and 0.069±0.031 (n=7) for EC-EC and SMC-SMC pairs, respectively). By contrast, the ECRs of heterocellular cells were significantly different when a current pulse (1.5 nA, 2s) was injected into EC and SMC respectively (0.072±0.025 for EC; 0.003±0.001 for SMC, n=5, P<0.01). The putative gap junction blocker 18ß-glycyrrhetinic acid significantly attenuated electrical coupling in both homocellular and heterocellular forms. The results suggest that homocellular GJs within SMCs or ECs are well coordinated but myoendothelial couplings between ECs and SMCs are unidirectional.

Inhibition of KIR2.1 decreases pulmonary artery smooth muscle cell proliferation and migration.

The investigation of effective therapeutic drugs for pulmonary hypertension (PH) is critical. KIR2.1 plays crucial roles in regulating cell proliferation and migration, and vascular remodeling. However, researchers have not yet clearly determined whether KIR2.1 participates in the proliferation and migration of pulmonary artery smooth muscle cells (PASMCs) and its role in pulmonary vascular remodeling (PVR) also remains elusive. The present study aimed to examine whether KIR2.1 alters PASMC proliferation and migration, and participates in PVR, as well as to explore its mechanisms of action. For the in vivo experiment, a PH model was established by intraperitoneally injecting Sprague­Dawley rats monocrotaline (MCT). Hematoxylin and eosin staining revealed evidence of PVR in the rats with PH. Immunofluorescence staining and western blot analysis revealed increased levels of the KIR2.1, osteopontin (OPN) and proliferating cell nuclear antigen (PCNA) proteins in pulmonary blood vessels and lung tissues following exposure to MCT, and the TGF­ß1/SMAD2/3 signaling pathway was activated. For the in vitro experiments, the KIR2.1 inhibitor, ML133, or the TGF­ß1/SMAD2/3 signaling pathway blocker, SB431542, were used to pre­treat human PASMCs (HPASMCs) for 24 h, and the cells were then treated with platelet­derived growth factor (PDGF)­BB for 24 h. Scratch and Transwell assays revealed that PDGF­BB promoted cell proliferation and migration. Immunofluorescence staining and western blot analysis demonstrated that PDGF­BB upregulated OPN and PCNA expression, and activated the TGF­ß1/SMAD2/3 signaling pathway. ML133 reversed the proliferation and migration induced by PDGF­BB, inhibited the expression of OPN and PCNA, inhibited the TGF­ß1/SMAD2/3 signaling pathway, and reduced the proliferation and migration of HPASMCs. SB431542 pre­treatment also reduced cell proliferation and migration; however, it did not affect KIR2.1 expression. On the whole, the results of the present study demonstrate that KIR2.1 regulates the TGF­ß1/SMAD2/3 signaling pathway and the expression of OPN and PCNA proteins, thereby regulating the proliferation and migration of PASMCs and participating in PVR.

[18ß-glycyrrhetinic acid inhibits outward current of vascular smooth muscle cells of arterioles].

The aim of the present study was to investigate the effect of 18ß-glycyrrhetinic acid (18ßGA) on the membrane current of vascular smooth muscle cells (VSMCs) in arteriole. Guinea pig anterior inferior cerebellar artery (AICA) and mesenteric artery (MA) were isolated, and single VSMCs were harvested using digestion with papain and collagenase IA. Outward currents of the VSMCs were recorded by whole-cell patch clamp technique. Results were shown as below: (1) 1 mmol/L 4-AP and 1 mmol/L TEA both could partially inhibit the whole-cell current of VSMCs in arterioles. (2) 18ßGA inhibited the outward current of VSMCs in a concentration-dependent manner. The inhibitory rates of 10, 30 and 100 µmol/L 18ßGA on the membrane current of VSMCs (+40 mV) were (25.3 ± 7.1)%, (43.1 ± 10.4)% and (68.4 ± 3.9)% respectively in AICA, and (13.2 ± 5.6)%, (34.2 ± 4.0)% and (59.3 ± 7.3)% respectively in MA. There was no significant difference between the inhibitory effects of 18ßGA on AICA and MA. 18ßGA also inhibited the outward current of VSMCs in a voltage-dependent manner. 18ßGA induced a more pronounced inhibition of the outward current from 0 to +40 mV, especially at +40 mV. (3) With the pretreatment of 10 mmol/L TEA, the inhibitory effect of 18ßGA on the membrane current of VSMCs was significantly abolished. These results suggest that the outward current of VSMCs in arterioles is mediated by voltage-dependent K(+) channels (K(v)) and big conductance calcium-activated K(+) channels (BK(Ca)), which can be inhibited by 18ßGA in concentration- and voltage-dependent way.

High pulse pressure is a risk factor for prodromal Alzheimer's disease: a longitudinal study.

It has been increasingly evident that pulse pressure (PP) is associated with Alzheimer's disease (AD) but whether PP increases AD risk and the mechanism responsible for this association remains unclear. To investigate the effects of PP in the process of AD, we have evaluated the cross-sectional and longitudinal associations of PP with AD biomarkers, brain structure and cognition and have assessed the effect of PP on AD risk in a large sample (n= 1,375) from the Alzheimer's Disease Neuroimaging Initiative (ADNI). Multiple linear regression and mixed-model regression were employed in cross-sectional and longitudinal analyses respectively. Clinical disease progression was assessed using Cox proportional hazards models. High PP was associated with lower ß-amyloid 42 (Aß42) (P= .015), and higher total tau (T-tau) (P= .011), phosphorylated tau (P-tau) (P= .003), T-tau/Aß42 (P= .004) and P-tau/Aß42 (P = .001), as well as heavier cortical amyloid-beta burden (P= .011). Longitudinally, baseline high PP was significantly associated with hippocampal atrophy (P= .039), entorhinal atrophy (P= .031) and worse memory performance (P= .058). Baseline high PP showed more rapid progression than those with normal PP (P <.001). These results suggest PP elevation could increase AD risk, which may be driven by amyloid plaques and subclinical neurodegeneration.

Upregulation of Nav1.7 by endogenous hydrogen sulfide contributes to maintenance of neuropathic pain.

Nav1.7 is closely associated with neuropathic pain. Hydrogen sulfide (H2S) has recently been reported to be involved in numerous biological functions, and it has been shown that H2S can enhance the sodium current density, and inhibiting the endogenous production of H2S mediated by cystathionine ß­synthetase (CBS) using O­(carboxymethyl)hydroxylamine hemihydrochloride (AOAA) can significantly reduce the expression of Nav1.7 and thus the sodium current density in rat dorsal root ganglion (DRG) neurons. In the present study, it was shown that the fluorescence intensity of H2S was increased in a spared nerve injury (SNI) model and AOAA inhibited this increase. Nav1.7 is expressed in DRG neurons, and the expression of CBS and Nav1.7 were increased in DRG neurons 7, 14 and 21 days post­operation. AOAA inhibited the increase in the expression of CBS, phosphorylated (p)­MEK1/2, p­ERK1/2 and Nav1.7 induced by SNI, and U0126 (a MEK blocker) was able to inhibit the increase in p­MEK1/2, p­ERK1/2 and Nav1.7 expression. However, PF­04856264 did not inhibit the increase in CBS, p­MEK1/2, p­ERK1/2 or Nav1.7 expression induced by SNI surgery. The current density of Nav1.7 was significantly increased in the SNI model and administration of AOAA and U0126 both significantly decreased the density. In addition, AOAA, U0126 and PF­04856264 inhibited the decrease in rheobase, and the increase in action potential induced by SNI in DRG neurons. There was no significant difference in thermal withdrawal latency among each group. However, the time the animals spent with their paw lifted increased significantly following SNI, and the time the animals spent with their paw lifted decreased significantly following the administration of AOAA, U0126 and PF­04856264. In conclusion, these data show that Nav1.7 expression in DRG neurons is upregulated by CBS­derived endogenous H2S in an SNI model, contributing to the maintenance of neuropathic pain.

Expression and effect of sodium-potassium-chloride cotransporter on dorsal root ganglion neurons in a rat model of chronic constriction injury.

Sodium-potassium-chloride cotransporter 1 (NKCC1) and potassium-chloride cotransporter 2 (KCC2) are associated with the transmission of peripheral pain. We investigated whether the increase of NKCC1 and KCC2 is associated with peripheral pain transmission in dorsal root ganglion neurons. To this aim, rats with persistent hyperalgesia were randomly divided into four groups. Rats in the control group received no treatment, and the rat sciatic nerve was only exposed in the sham group. Rats in the chronic constriction injury group were established into chronic constriction injury models by ligating sciatic nerve and rats were given bumetanide, an inhibitor of NKCC1, based on chronic constriction injury modeling in the chronic constriction injury + bumetanide group. In the experiment measuring thermal withdrawal latency, bumetanide (15 mg/kg) was intravenously administered. In the patch clamp experiment, bumetanide (10 µg/µL) and acutely isolated dorsal root ganglion neurons (on day 14) were incubated for 1 hour, or bumetanide (5 µg/µL) was intrathecally injected. The Hargreaves test was conducted to detect changes in thermal hyperalgesia in rats. We found that the thermal withdrawal latency of rats was significantly decreased on days 7, 14, and 21 after model establishment. After intravenous injection of bumetanide, the reduction in thermal retraction latency caused by model establishment was significantly inhibited. Immunohistochemistry and western blot assay results revealed that the immune response and protein expression of NKCC1 in dorsal root ganglion neurons of the chronic constriction injury group increased significantly on days 7, 14, and 21 after model establishment. No immune response or protein expression of KCC2 was observed in dorsal root ganglion neurons before and after model establishment. The Cl- (chloride ion) fluorescent probe technique was used to evaluate the change of Cl- concentration in dorsal root ganglion neurons of chronic constriction injury model rats. We found that the relative optical density of N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (a Cl- fluorescent probe whose fluorescence intensity decreases as Cl- concentration increases) in the dorsal root ganglion neurons of the chronic constriction injury group was significantly decreased on days 7 and 14 after model establishment. The whole-cell patch clamp technique revealed that the resting potential and action potential frequency of dorsal root ganglion neurons increased, and the threshold and rheobase of action potentials decreased in the chronic constriction injury group on day 14 after model establishment. After bumetanide administration, the above indicators were significantly suppressed. These results confirm that CCI can induce abnormal overexpression of NKCC1, thereby increasing the Cl- concentration in dorsal root ganglion neurons; this then enhances the excitability of dorsal root ganglion neurons and ultimately promotes hyperalgesia and allodynia. In addition, bumetanide can achieve analgesic effects. All experiments were approved by the Institutional Ethics Review Board at the First Affiliated Hospital, College of Medicine, Shihezi University, China on February 22, 2017 (approval No. A2017-169-01).

Occurrence and source analysis of selected antidepressants and their metabolites in municipal wastewater and receiving surface water.

Seven commonly utilized antidepressants (amitriptyline, fluoxetine, sertraline, citalopram, paroxetine, venlafaxine, and bupropion) and three of their metabolites were detected in four wastewater treatment plants (WWTPs) and corresponding receiving waters including the mainstream and three of the tributaries of Huangpu River, Shanghai. The concentration levels of selected antidepressants in wastewater and receiving water were both at ng L-1 level, but those antidepressants that were found in wastewater were typically in a range of one and even two orders of magnitude higher than those that were present in the receiving water except for the concentration levels of paroxetine, norfluoxetine, and nortriptyline. Venlafaxine and its metabolite O-desmethylvenlafaxine were the primary compounds (reaching up to 132.04 and 173.68 ng L-1 as well as 3.03 and 4.53 ng L-1 in wastewater and receiving water, respectively). Based on the mass loadings of selected antidepressants in four WWTPs, the mass loading of sertraline in effluent for this study was at the highest level when compared to other countries. The important finding was that the sampling sites H11 and H13 were much more likely to be polluted by side-pollution sources (the accumulation areas of Grade A of Class-three hospitals and pharmaceutical factories) than point pollution sources (WWTP 1 and WWTP2) through analysis of Spearman correlation. The results have shown that the RQs for these antidepressants were less than 0.1 except for the RQ of venlafaxine in an effluent for mollusks, which was less than 1. This indicated that the concentration levels of antidepressants would not pose a high risk for aquatic organisms, but sustained attention should still be paid.

Association of connexin gene polymorphism with essential hypertension in Kazak and Han Chinese in Xinjiang, China.

Essential hypertension (EH) is affected by both genetic and environmental factors. The polymorphism of connexin (Cx) genes is found associated with the development of hypertension. However, the association of the polymorphism of Cxs with EH has not been investigated. This study aimed to investigate the association of the polymorphism of connexin (Cx) genes Cx37, Cx40, and Cx43 with EH in Kazak and Han Chinese in Xinjiang, China. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) were used to analyze the polymorphism of Cx genes in Kazak and Han EH patients as well as their normotensive controls. The results showed that there were no significant differences in the frequencies of different three genotypes (A/A, A/G, and G/G) and A and G alleles of Cx40 rs35594137 and rs11552588 between EH patients and normotensive controls. However, in Kazak EH patients, the frequencies of three genotypes (A/A, A/G, and G/G) of Cx37 rs1630310 were 24.8%, 47.2% and 28.0%, respectively, which were significantly different from those in Han EH patients. In Han EH patients, the frequencies of the three genotypes (C/C, C/G and G/G) of Cx43 rs1925223 were 6.4%, 35.6% and 58.0%, respectively. Frequencies of the other four genotypes had no statistical differences among Kazak and Han EH patients and their normotensive controls. These results suggest polymorphisms of Cx37 rs1630310 and Cx43 rs1925223 genes may be associated with the pathogenesis of EH. Carrying Cx37 rs1630310-A or Cx43 rs1925223-G genotypes may protect against the development of EH.

Enhanced expression of Cx43 and gap junction communication in vascular smooth muscle cells of spontaneously hypertensive rats.

Niflumic acid (NFA) is a novel gap junction (GJ) inhibitor. The aim of the present study was to investigate the effect of NFA on GJ communication and the expression of connexin (Cx) in vascular smooth muscle cells (VSMCs) of mesenteric arterioles of spontaneously hypertensive rats (SHR). Whole­cell patch clamp recording demonstrated that NFA at 1x10­4 M significantly inhibited the inward current and its effect was reversible. The time for charging and discharging of cell membrane capacitance (Cinput) reduced from 9.73 to 0.48 ms (P<0.05; n=6). Pressure myograph measurement showed that NFA at 3x10­4 M fully neutralized the contraction caused by phenylephrine. The relaxation responses of normotensive control Wistar Kyoto (WKY) rats were significantly higher, compared with those of the SHRs (P<0.05; n=6). Western blot and reverse transcription­quantitative polymerase chain reaction analyses showed that the mRNA and protein expression levels of Cx43 of the third­level branch of mesenteric arterioles of the SHRs and WKY rats were higher, compared with those of the first­level branch. The mRNA and protein expression levels of Cx43 of the primary and third­level branches of the mesenteric arterioles in the SHRs were higher, compared with those in the WKY rats (P<0.05; n=6). The mRNA levels of Cx43 in the mesenteric arterioles were significantly downregulated by NFA in a concentration­dependent manner (P<0.01; n=6). The protein levels of Cx43 in primary cultured VSMCs isolated from the mesenteric arterioles were also significantly downregulated by NFA in a concentration­dependent manner (P<0.01; n=6). These results showed that the vasorelaxatory effects of GJ inhibitors were reduced in the SHRs, which was associated with a higher protein expression level of Cx43 in the mesenteric arterioles of the SHRs. NFA also relaxed the mesenteric arterioles by reducing the expression of Cx43, which decreased blood pressure. Therefore, regulation of the expression of GJs may be a therapeutic target for the treatment of hypertension.

NSAIDs modulate GABA-activated currents via Ca2+-activated Cl- channels in rat dorsal root ganglion neurons.

The ability of non-steroidal anti-inflammatory drugs (NSAIDs) to modulate γ-aminobutyrate (GABA)-activated currents via Ca2+-activated Cl- channels in rat dorsal root ganglion neurons (DRG), was examined in the present study. During the preparation of DRG neurons harvested from Sprague-Dawley rats, the whole-cell recording technique was used to record the effect of NSAIDs on GABA-activated inward currents, and the expression levels of the TMEM16A and TMEM16B subunits were revealed. In the event that DRG neurons were pre-incubated for 20 sec with niflumic acid (NFA) and 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) prior to the administration of GABA, the GABA-induced inward currents were diminished markedly in the majority of neurons examined (96.3%). The inward currents induced by 100 µmol/l GABA were attenuated by (0±0.09%; neurons = 4), (5.32±3.51%; neurons = 6), (21.3±4.00%; neurons = 5), (33.8±5.20%; neurons = 17), (52.2±5.10%; neurons = 4) and (61.1±4.12%; neurons = 12) by 0.1, 1, 3, 10, 30 and 100 µmol/l NFA, respectively. The inward currents induced by 100 µmol/l GABA were attenuated by (13.8±6%; neurons = 6), (23.2±14.7%; neurons = 6) and (29.7±9.1%; neurons = 9) by 3, 10 and 30 µmol/l NPPB, respectively. NFA and NPPB dose-dependently inhibited GABA-activated currents with half maximal inhibitory concentration (IC50) values of 6.7 and 11 µmol/l, respectively. The inhibitory effect of 100 µmol/l NFA on the GABA-evoked inward current were also strongly inhibited by nitrendipine (NTDP; an L-type calcium channel blocker), 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis (a highly selective calcium chelating reagent), caffeine (a widely available Ca2+ consuming drug) and calcium-free extracellular fluid, in a concentration-dependent manner. Immunofluorescent staining indicated that TMEM16A and TMEM16B expression was widely distributed in DRG neurons. The results suggest that NSAIDs may be able to regulate Ca2+-activated chloride channels to reduce GABAA receptor-mediated inward currents in DRGs.

Vaccination with Bivalent DNA Vaccine of α1-Giardin and CWP2 Delivered by Attenuated Salmonella typhimurium Reduces Trophozoites and Cysts in the Feces of Mice Infected with Giardia lamblia.

BACKGROUND: Giardia lamblia is one of the most common infectious protozoans in human that may cause diarrhea in travelers. Searching for antigens that induced effectively protective immunity has become a key point in the development of vaccine against giardiasis. METHODOLOGY/PRINCIPAL FINDINGS: Mice vaccinated with G. lamblia trophozozite-specific α1-giardin DNA vaccine delivered orally by attenuated Salmonella typhimurium SL7027 elicited 74.2% trophozoite reduction, but only 28% reduction in cyst shedding compared with PBS buffer control. Oral vaccination with Salmonella-delivered cyst-specific CWP2 DNA produced 89% reduction in cysts shedding in feces of vaccinated mice. Significantly, the mice vaccinated with Salmonella-delivered bivalent α1-giardin and CWP2 DNA vaccines produced significant reduction in both trophozoite (79%) and cyst (93%) in feces of vaccinated mice. This parasite reduction is associated with the strong local mucosal IgA secretion and the IgG2a-dominant systemic immune responses in vaccinated mice. CONCLUSIONS: The results demonstrate that bivalent vaccines targeting α1-giardin and CWP2 can protect mice against the colonization of Giardia trophozoite and block the transformation of cyst in host at the same time, and can be used to prevent Giardia infection and block the transmission of giardiasis.

[The characteristics of resting membrane potential on smooth muscle cells and endothelial cells in guinea pigs cochlea spiral artery].

OBJECTIVE: A variety of inner ear disease is related to microcirculation disturbance of inner ear, but smooth muscle cells (SMC) and endothelial cells (EC) of the spiral modiolar artery (SMA), which is the main blood supply to the inner ear, physiological feature is not very clear. METHODS: In this study, two-intracellular microelectrode recording technique and cell staining techniques to study the SMC and EC resting membrane potential characteristics and communication links between cells of SMA. RESULTS: Study found that SMC and EC have high and low resting membrane potential state, two state of the resting membrane potential of cells to ACh and high K+ response is completely different. The different types of cells, EC-EC, SMC-SMC and SMC-EC, can simultaneously record by two-microelectrode, two cell resting membrane potential can also be a double-high RP, double-low RP and one high- and one low- RP. Experiment recorded in one high- and one low- RP are the SMC-EC types, and ECs initial membrane potential are high potential, SMCs membrane potential are low initial potential. The double-high and double-low RP can be SMC-SMC or EC-EC or SMC-EC types. CONCLUSION: The results show that SMC and EC in the 0.3 - 0.5 mm range, similar type of cells have very good communication, can function together to maintain good and consistent, heterogeneous cell performance is more different.

[The distribution and mechanism of coronary arteriole cell resting membrane potential in guinea pigs].

OBJECTIVE: To investigate the distribution and mechanism of coronary arteriole (CA) cell resting membrane potential (RP) in guinea pigs. METHODS: Cell RP was recorded by intracellular microelectrode in isolated guinea pig coronary arteriole (diameter < 100 microm). RESULTS: (1) Experiments were carried out in 112 cells with a mean RP of (-65 +/- 4.2)mV, the distribution of coronary arteriole cell RP fitted by Gaussian function was bimodal, one peak was -43 mV termed high RP, the other was -74 mV termed low RP. 10 mmol/L K+ and 3 micromol/ L acetylcholine(ACh) induced hyperpolarization in high-RP cells with (-7.4 +/- 0.87) mV (n = 13) and (-15 +/- 1.24) mV (n = 16) respectively, and induced depolarization in low-RP cells with (9.6 +/- 1.2) mV (n = 23) and (8.7 +/- 0.69) mV (n = 15) respectively. (2) The inward rectifier K+ channel (K(ir)) blocker Ba2+ caused concentration-dependent depolarization in low-RP cells with an EC50 of 120 micromol/L 100 micromol/L Ba2+ or higher could shift low-RP cells to high-RP state, the response of these cells to high K+ and ACh became a hyperpolarization. CONCLUSION: The distribution of coronary vascular cell RP is bimodal, high K+ and ACh induce different responses in low and high RP cells. The two RP states are exchangeable mainly due to all-or-none conductance changes of K(ir).