Therapeutic potential of the secreted Kazal-type serine protease inhibitor SPINK4 in colitis.

Mucus injury associated with goblet cell (GC) depletion constitutes an early event in inflammatory bowel disease (IBD). Using single-cell sequencing to detect critical events in mucus dysfunction, we discover that the Kazal-type serine protease inhibitor SPINK4 is dynamically regulated in colitic intestine in parallel with disease activities. Under chemically induced colitic conditions, the grim status in Spink4-conditional knockout mice is successfully rescued by recombinant murine SPINK4. Notably, its therapeutic potential is synergistic with existing TNF-α inhibitor infliximab in colitis treatment. Mechanistically, SPINK4 promotes GC differentiation using a Kazal-like motif to modulate EGFR-Wnt/ß-catenin and -Hippo pathways. Microbiota-derived diacylated lipoprotein Pam2CSK4 triggers SPINK4 production. We also show that monitoring SPINK4 in circulation is a reliable noninvasive technique to distinguish IBD patients from healthy controls and assess disease activity. Thus, SPINK4 serves as a serologic biomarker of IBD and has therapeutic potential for colitis via intrinsic EGFR activation in intestinal homeostasis.

ICG-Dimeric Her2-Specific Affibody Conjugates for Tumor Imaging and Photothermal Therapy for Her2-Positive Tumors.

Human epidermal growth factor receptor 2 (Her2) is abundantly expressed in various solid tumors. The Her2-specific Affibody (ZHer2:2891) has been clinically tested in patients with Her2-positive breast cancer and is regarded as an ideal drug carrier for tumor diagnosis and targeted treatment. Indocyanine green (ICG) can be used as a photosensitizer for photothermal therapy (PTT), in addition to fluorescent dyes for tumor imaging. In this study, a dimeric Her2-specific Affibody (ZHer2) based on ZHer2:2891 was prepared using the E. coli expression system and then coupled to ICG through an N-hydroxysuccinimide (NHS) ester reactive group to construct a novel bifunctional protein drug (named ICG-ZHer2) for tumor diagnosis and PTT. In vitro, ICG-ZHer2-mediated PTT selectively and efficiently killed Her2-positive BT-474 and SKOV-3 tumor cells rather than Her2-negative HeLa tumor cells. In vivo, ICG-ZHer2 specifically accumulated in Her2-positive SKOV-3 tumor grafts rather than Her2-negative HeLa tumor grafts; high-contrast tumor optical images were obtained. However, Her2-negative HeLa tumor grafts were not detected. More importantly, ICG-ZHer2-mediated PTT exhibited a significantly enhanced antitumor effect in mice bearing SKOV-3 tumor grafts owing to the good photothermal properties of ICG-ZHer2. Of note, ICG-ZHer2 did not exhibit acute toxicity in mice during short-term treatment. Overall, our findings indicate that ICG-ZHer2 is a promising bifunctional drug for Her2-positive tumor diagnosis and PTT.

Her3-specific affibody mediated tumor targeting delivery of ICG enhanced the photothermal therapy against Her3-positive tumors.

Photothermal therapy (PTT), mediated by tumor-targeted drug delivery of indocyanine green (ICG), is a promising strategy for cancer therapy. Human epidermal growth factor receptor 3 (Her3) is highly expressed in several solid tumors and is an ideal target for tumor diagnosis and therapy. This study prepared a Her3-specific dimeric affibody (ZHer3) using an Escherichia coli expression system. The affibody could bind explicitly to Her3-positive MCF7 and LS174T cells, rather than to Her3-negative SKOV-3 cells in vitro. ICG was coupled with the ZHer3 affibody (ICG-ZHer3) through an N-hydroxysuccinimide (NHS) ester reactive group for tumor-targeted delivery. As expected, Her3-positive cells were selectively and efficiently killed by ICG-ZHer3-mediated PTT in vitro. In vivo, ICG-ZHer3 preferentially accumulated in Her3-positive LS174T tumor grafts because of the tumor-targeting ability of the ZHer3 affibody. As a result of the local generation of cytotoxic reactive oxygen species and hyperthermia, the growth rates of LS174T tumor grafts were significantly inhibited by ICG-ZHer3-mediated PTT, and ICG-ZHer3 showed good safety performance during short-term treatment. In conclusion, these results demonstrated that ICG-ZHer3 is a promising photosensitizer for PTT against Her3-positive tumors.

The role of Gα protein signaling in the membrane estrogen receptor-mediated signaling.

Estrogens exert rapid, extranuclear effects by their action on the plasma membrane estrogen receptors (mERs). Gα protein associated with the cell membrane is involved in many important processes regulated by estrogens. However, the Gα's role in the mER-mediated signaling and the signaling pathways involved are poorly understood. This review aims to outline the Gα's role in the mER-mediated signaling. Immunoblotting, immunofluorescence, co-immunoprecipitation, and RNA interference were carried out using vascular endothelial cells (ECs) and human breast carcinoma cell lines as experimental models. Electrophysiology and immunocytochemistry were carried out using guinea pigs as animal models. Recent advances suggest that the signaling of mERα through Gα is required for vascular EC migration or endothelial H2S release, while Gα13 is involved in estrogen-induced breast cancer cell invasion. Besides, the Gαq-coupled PLC-PKC-PKA pathway is critical for the neural regulation of energy homeostasis. This review summarizes the contributions of Gα to mER-mediated signaling, including cardiovascular protection, breast cancer metastasis, neural regulation of homeostatic functions, and osteogenesis.

Hydrogen Gas: A Novel Type of Antioxidant in Modulating Sexual Organs Homeostasis.

Sex is a science of cutting edge but bathed in mystery. Coitus or sexual intercourse, which is at the core of sexual activities, requires healthy and functioning vessels to supply the pelvic region, thus contributing to clitoris erection and vaginal lubrication in female and penile erection in male. It is well known that nitric oxide (NO) is the main gas mediator of penile and clitoris erection. In addition, the lightest and diffusible gas molecule hydrogen (H2) has been shown to improve erectile dysfunction (ED), testis injuries, sperm motility in male, preserve ovarian function, protect against uterine inflammation, preeclampsia, and breast cancer in female. Mechanistically, H2 has strong abilities to attenuate excessive oxidative stress by selectively reducing cytotoxic oxygen radicals, modulate immunity and inflammation, and inhibit injuries-induced cell death. Therefore, H2 is a novel bioactive gas molecule involved in modulating sexual organs homeostasis.

Helium Protects Against Lipopolysaccharide-Induced Cardiac Dysfunction in Mice via Suppressing Toll-Like Receptor 4-Nuclear Factor κB-Tumor Necrosis Factor-Alpha/ Interleukin-18 Signaling.

The nonanesthetic noble gas helium (He) can protect many organs against ischemia and reperfusion injury, such as liver and heart. However, the role of He on cardiac dysfunction during sepsis is not clear. In this study, we established a lipopolysaccharide (LPS)-induced cardiac dysfunction mouse model to examine the influence of He on the impaired cardiac function, and further investigated the possible innate immune mechanisms that may be involved. LPS induced left ventricular dysfunction and cavity enlargement, as indicated by decreased percent ejection fraction, percent fractional shortening, left ventricular anterior wall thickness in systole, and left ventricular posterior wall thickness in systole, while increased left ventricular end-systolic diameter and left ventricular end-systolic volume. He improved the impaired left ventricular function and cavity enlargement in a dose-dependent manner, and it was beneficial at 1.0 mL/100 g. Mechanistically, He inhibited toll-like receptor 4 (TLR4) expression, reduced the phosphorylation of nuclear factor κB (NF-κB), and subsequently alleviated tumor necrosis factor-alpha (TNF-α) and interleukin-18 (IL-18) expression in heart. Therefore, He protects against LPS-induced cardiac dysfunction in mice partially via inhibiting myocardial TLR4-NF-κB-TNF-α/IL-18 signaling.

The FTO/miR-181b-3p/ARL5B signaling pathway regulates cell migration and invasion in breast cancer.

BACKGROUND: N6-methyladenosine (m6 A) RNA modification has been demonstrated to be a significant regulatory process in the progression of various tumors, including breast cancer. Fat mass and obesity-associated (FTO) enzyme, initially known as the obesity-related protein, is the first identified m6 A demethylase. However, the relationship between FTO and breast cancer remains controversial. In this study, we aimed to elucidate the role and clinical significance of FTO in breast cancer and to explore the underlying mechanism. METHODS: We first investigated the expression of FTO in breast cancer cell lines and tissues by quantitative reverse transcription-PCR (qRT-PCR), Western blotting, and immunohistochemistry. Wound healing assay and Transwell assay were performed to determine the migration and invasion abilities of SKBR3 and MDA-MB453 cells with either knockdown or overexpression of FTO. RNA sequencing (RNA-seq) was conducted to decipher the downstream targets of FTO. qRT-PCR, luciferase reporter assay, and Western blotting were employed to confirm the existence of the FTO/miR-181b-3p/ARL5B axis. The biological function of ADP ribosylation factor like GTPase 5B (ARL5B) in breast cancer cells was evaluated by wound healing assay and Transwell invasion assay. RESULTS: High FTO expression was observed in human epidermal growth factor receptor 2 (HER2)-positive breast cancer, predicting advanced progression (tumor size [P < 0.001], nuclear grade [P = 0.001], peritumoral lymphovascular invasion [P < 0.001), lymph node metastasis [P = 0.002], and TNM stage [P = 0.001]) and poor prognosis. Moreover, FTO promoted cell invasion and migration in vitro. Mechanistically, RNA-seq and further confirmation studies suggested that FTO up-regulated ARL5B by inhibiting miR-181b-3p. We further verified that ARL5B also displayed carcinogenic activity in breast cancer cells. CONCLUSION: Our work demonstrated the carcinogenic activity of FTO in promoting the invasion and migration of breast cancer cells via the FTO/miR-181b-3p/ARL5B signaling pathway.

Prognostic role of GPER/Ezrin in triple-negative breast cancer is associated with menopausal status.

The role of G protein-coupled estrogen receptor 1 (GPER) signaling, including promotion of Ezrin phosphorylation (which could be activated by estrogen), has not yet been clearly identified in triple-negative breast cancer (TNBC). This study aimed to evaluate the prognostic value of GPER and Ezrin in TNBC patients. Clinicopathologic features including age, menopausal status, tumor size, nuclear grade, lymph node metastasis, AJCC TNM stage, and ER, PR and HER-2 expression were evaluated from 249 TNBC cases. Immunohistochemical staining of GPER and Ezrin was performed on TNBC pathological sections. Kaplan-Meier analyses, as well as logistic regressive and Cox regression model tests were applied to evaluate the prognostic significance between different subgroups. Compared to the GPER-low group, the GPER-high group exhibited higher TNM staging (P = 0.021), more death (P < 0.001), relapse (P < 0.001) and distant events (P < 0.001). Kaplan-Meier analysis showed that GPER-high patients had a decreased OS (P < 0.001), PFS (P < 0.001), LRFS (P < 0.001) and DDFS (P < 0.001) than GPER-low patients. However, these differences in prognosis were not statistically significant in post-menopausal patients (OS, P = 0.8617; PFS, P = 0.1905; LRFS, P = 0.4378; DDFS, P = 0.2538). There was a significant positive correlation between GPER and Ezrin expression level (R = 0.508, P < 0.001) and the effect of Ezrin on survival prognosis corresponded with GPER. Moreover, a multivariable analysis confirmed that GPER and Ezrin level were both significantly associated with poor DDFS (HR: 0.346, 95% CI 0.182-0.658, P = 0.001; HR: 0.320, 95% CI 0.162-0.631, P = 0.001). Thus, overexpression of GPER and Ezrin may contribute to aggressive behavior and indicate unfavorable prognosis in TNBC; this may correspond to an individual's estrogen levels.

MET mutation causes muscular dysplasia and arthrogryposis.

Arthrogryposis is a group of phenotypically and genetically heterogeneous disorders characterized by congenital contractures of two or more parts of the body; the pathogenesis and the causative genes of arthrogryposis remain undetermined. We examined a four-generation arthrogryposis pedigree characterized by camptodactyly, limited forearm supination, and loss of myofibers in the forearms and hands. By using whole-exome sequencing, we confirmed MET p.Y1234C mutation to be responsible for arthrogryposis in this pedigree. MET p.Y1234C mutation caused the failure of activation of MET tyrosine kinase. A Met p.Y1232C mutant mouse model was established. The phenotypes of homozygous mice included embryonic lethality and complete loss of muscles that originated from migratory precursors. Heterozygous mice were born alive and showed reduction of the number of myofibers in both appendicular and axial muscles. Defective migration of muscle progenitor cells and impaired proliferation of secondary myoblasts were proven to be responsible for the skeletal muscle dysplasia of mutant mice. Overall, our study shows MET to be a causative gene of arthrogryposis and MET mutation could cause skeletal muscle dysplasia in human beings.

Estrogen promotes the onset and development of idiopathic scoliosis via disproportionate endochondral ossification of the anterior and posterior column in a bipedal rat model.

This study aimed to verify the effects of estrogen on the onset and development of adolescent idiopathic scoliosis and the mechanisms associated with these effects by constructing a pubescent bipedal rat model. Experiments were conducted to investigate whether scoliosis progression was prevented by a Triptorelin treatment. One hundred twenty bipedal rats were divided into female, OVX (ovariectomy), OVX + E2, Triptorelin, sham, and male groups. According to a spinal radiographic analysis, the scoliosis rates and curve severity of the female and OVX + E2 groups were higher than those in the OVX, Triptorelin, and male groups. The measurements obtained from the sagittal plane of thoracic vertebrae CT confirmed a relatively slower growth of the anterior elements and a faster growth of the posterior elements between T11 and T13 in the female and OVX + E2 groups than in the OVX and Triptorelin groups. Histomorphometry and immunohistochemistry revealed a significantly longer hypertrophic zone of the vertebral cartilage growth plates that expressed more type X collagen and less type II collagen in the OVX and Triptorelin groups than in the female and OVX + E2 groups. Ki67 immunostaining confirmed an increase in the proliferation of vertebral growth plate chondrocytes in the OVX group compared with the female and OVX + E2 groups. In conclusion, estrogen obviously increased the incidence of scoliosis and curve severity in pubescent bipedal rats. The underlying mechanism may be a loss of coupling of the endochondral ossification between the anterior and posterior columns. Triptorelin decreased the incidence of scoliosis and curve magnitudes in bipedal female rats.

17ß-estradiol upregulates striatin protein levels via Akt pathway in human umbilical vein endothelial cells.

17ß-estradiol (E2) has been shown to have beneficial effects on the cardiovascular system. We previously demonstrated that E2 increases striatin levels and inhibits migration in vascular smooth muscle cells. The objective of the present study was to investigate the effects of E2 on the regulation of striatin expression in human umbilical vein endothelial cells (HUVECs). We demonstrated that E2 increased striatin protein expression in a dose- and time-dependent manner in HUVECs. Pretreatment with ICI 182780 or the phosphatidylinositol-3 kinase inhibitor, wortmannin, abolished E2-mediated upregulation of striatin protein expression. Treatment with E2 resulted in Akt phosphorylation in a time-dependent manner. Moreover, silencing striatin significantly inhibited HUVEC migration, while striatin overexpression significantly promoted HUVEC migration. Finally, E2 enhanced HUVEC migration, which was inhibited by silencing striatin. In conclusion, our results demonstrated that E2-mediated upregulation of striatin promotes cell migration in HUVECs.

Dual novel mutations in SLC26A2 in two siblings with multiple epiphyseal dysplasia 4 from a Chinese family: a case report.

BACKGROUND: Multiple epiphyseal dysplasia (MED) is a heterogeneous genetic condition characterized by variable phenotypes, such as short stature (mild to moderate), joint deformities, abnormal gait, scoliosis, and brachydactyly. Recessive mutations in the SLC26A2 gene cause a phenotype of multiple epiphyseal dysplasia-4 (MED-4). In the present study, we identified novel compound heterozygous mutations in the SLC26A2 gene in a Chinese family with two affected sibs with MED-4. CASE PRESENTATION: Radiographs revealed hip dysplasia, brachydactyly and scoliosis in patient 1. Radiological examinations in patient 2 also showed hip dysplasia recently. Both of them were diagnosed with MED-4. SLC26A2 c.824 T > C and SLC26A2 c.1198C > T were identified in two siblings in this family, which were inherited from both parents, one mutation from each. CONCLUSIONS: This is the first Chinese MED-4 family attributed to SLC26A2 mutations, and these results show that these novel compound heterozygous mutations in SLC26A2 contribute to MED-4.

Decreased expression of protocadherin 20 is associated with poor prognosis in hepatocellular carcinoma.

Recently, protocadherin 20 has been reported as a tumor suppressor gene in hepatocellular carcinoma (HCC); however, the prognostic value of protocadherin 20 in HCC remains unclear. Hence, the purpose of this study was to investigate the clinical and prognostic values of protocadherin 20 in HCC patients. The expression of protocadherin 20 was assessed by quantitative real-time polymerase chain reaction, western blot, and immunohistochemistry. Kaplan-Meier curves were created to calculate the overall survival of the patients, and Cox regression models were used to identify the risk factors associated with prognosis. Of 317 primary HCC patients, decreased expression of protocadherin 20 was observed in 184 (58.0%) patients (P < 0.001). Reduced protocadherin 20 protein expression correlated with portal hypertension, poor tumor differentiation, advanced Okuda stage, and Cancer of the Liver Italian Program score (all P < 0.05). Low protocadherin 20 expression was an independent risk factor for mortality (P = 0.018). Furthermore, in our newly developed simple risk score based on protocadherin 20, patients with total score > 1.11 showed significantly poorer outcome; and the predictive value of the score was better than the Barcelona Clinic Liver Cancer stage, Okuda stage, and Child-Pugh classification (Harrell's concordance index = 0.614). Taken together, these findings suggest that protocadherin 20 may represent a novel prognostic biomarker for HCC patients.

Oestrogen exerts anti-inflammation via p38 MAPK/NF-κB cascade in adipocytes.

BACKGROUND: Oestrogen has anti-inflammatory property in obesity. However, the mechanism is still not defined. OBJECTIVE: To investigate the effect of oestrogen on LPS-induced monocyte chemoattractant protein-1 (MCP-1) production in adipocytes. METHODS: Lipopolysaccharides (LPS) was used to imitate inflammatory responses and monocyte chemotactic protein-1 (MCP-1) was selected as an inflammatory marker to observe. 17ß-Estradiol (E2), SB203580 (SB), pyrrolidine dithiocarbamate (PDTC), pertussis toxin (PTX), wortmannin (WM), p65 siRNA and p38 MAPK siRNA were pre-treated respectively or together in LPS-induced MCP-1. Then p38 MAPK and NF-κB cascade were silenced successively to observe the change of each other. Lastly, oestrogen receptor (ER) α agonist, ERß agonist and ER antagonist were utilised. RESULTS: LPS-induced MCP-1 largely impaired by pre-treatment with E2, SB, PDTC or silencing NF-κB subunit. E2 inhibited LPS-induced MCP-1 in a time- and dose-dependent manner, which was related to the suppression of p65 translocation to nucleus. Furthermore, LPS rapidly activated p38 MAPK, while E2 markedly inhibited this activation. It markedly attenuated LPS-stimulated p65 translocation to nucleus and MCP-1 production by transfecting with p38 MAPK siRNA or using p38 MAPK inhibitor. The oestrogen's inhibitory effect was mimicked by the ERα agonist, but not by the ERß agonist. The inhibition of E2 on p38 MAPK phosphorylation was prevented by ER antagonist. CONCLUSIONS: E2 inhibits LPS-stimulated MCP-1 in adipocytes. This effect is related to the inhibition of p38 MAPK/NF-κB cascade, and ERα appears to be the dominant ER subtype in these events.

Curcumin Ameliorates Reserpine-Induced Gastrointestinal Mucosal Lesions Through Inhibiting IκB-α/NF-κB Pathway and Regulating Expression of Vasoactive Intestinal Peptide and Gastrin in Rats.

The objective of our study was to investigate whether curcumin protects against reserpine-induced gastrointestinal mucosal lesions (GMLs) in rats and to explore the mechanism of curcumin's action. Sprague-Dawley rats were randomly divided into four groups: control group, reserpine-treated group, reserpine treatment group with curcumin at high dose (200 mg/kg), and reserpine treatment group with curcumin at low dose (100 mg/kg). Rats in reserpine-treated group were induced by intraperitoneally administered reserpine (0.5 mg/kg) for 28 days. TUNEL staining and hematoxylin and eosin staining were used to evaluate the apoptotic cells and morphologic changes. In addition, to explore the mechanism of curcumin in protecting GMLs, we used serum of experimental rats to assess the level of vasoactive intestinal peptide (VIP), gastrin, interleukin-6, interleukin-10, tumor necrosis factor-α and interferon-γ by ELISA and radioimmunoassay. The protein levels of NF-κB, p-IκB-α, IκB-α, Bcl-2, Bax, and cleaved-caspase-3 were examined by western blot analysis. Data were analyzed with SPSS 19.0 software package. Curcumin treatment prevented tissue damage and cell death in the reserpine-treated rats and effectively decreased inflammatory response and balanced the expression of VIP and gastrin in the reserpine-treated rats. NF-κB, p-IκB-α, Bax, and cleaved-caspase-3 were increased in the reserpine group, but the curcumin high-dose group inhibited them. Curcumin can target the IκB-α/NF-κB pathway to inhibit inflammatory response and regulate the level of VIP and gastrin in reserpine-induced GML rats.

17ß-Estradiol inhibits vascular smooth muscle cell migration via up-regulation of striatin protein.

Striatin, an estrogen receptor (ER)-interacting protein, plays an important role in estrogen's nongenomic actions in vascular endothelial cells. However, the role of striatin in VSMCs is unknown. Here, we investigated the role of striatin in estrogen-regulated VSMCs migration. 17ß-Estradiol (E2) at 10 nM largely inhibited VSMCs migration, which was reversed by the silencing of striatin expression. E2 increased striatin protein expression in a dose- and time-dependent manner. ERα agonist PPT, but not ERß agonist DPN, mimicked the regulatory effect of E2. The regulatory effect of E2 on striatin protein expression was blocked by the pure ER antagonist ICI 182,780 or the mitogen-activated protein kinase inhibitor PD98059, but not by the phosphatidylinositol-3 kinase inhibitor wortmannin or Src inhibitor PP2, suggesting that E2 increased striatin protein expression via extracellular-signal regulated kinase 1/2 (ERK1/2). E2 resulted in phosphorylation of ERK1/2 in a time-dependent manner. The silencing of ERK1/2 largely abolished E2-enhanced striatin expression. Finally, the inhibitory effect of E2 on VSMC migration was reversed by ICI 182,780 or PD98059. Taken together, our results indicate that E2 inhibits VSMC migration by increasing striatin expression via ERα to ERK1/2 pathway, which maybe helpful to understand estrogen's anti-atherogenic effect in VSMCs.

Protective effects of scutellarin on type II diabetes mellitus-induced testicular damages related to reactive oxygen species/Bcl-2/Bax and reactive oxygen species/microcirculation/staving pathway in diabetic rat.

The goal of our study is to evaluate the effect of Scutellarin on type II diabetes-induced testicular disorder and show the mechanism of Scutellarin's action. We used streptozotocin and high-fat diet to establish type II diabetic rat model. TUNEL and haematoxylin and eosin staining were used to evaluate the testicular apoptotic cells and morphologic changes. Immunohistochemical staining was used to measure the expression level of vascular endothelial growth factor and blood vessel density in testes. Oxidative stress in testes and epididymis was tested by fluorescence spectrophotometer and ELISA. The expression of Bcl-2/Bax and blood flow rate in testicular vessels were measured by western blot and Doppler. Our results for the first time showed that hyperglycemia induced apoptotic cells and morphologic impairments in testes of rats, while administration of Scutellarin can significantly inhibit these damages. This effect of Scutellarin is controlled by two apoptotic triggers: ROS/Bcl-2/Bax and ROS/microcirculation/starving pathway.

2-methoxyestradiol induces vasodilation by stimulating NO release via PPARγ/PI3K/Akt pathway.

The endogenous estradiol metabolite 2-methoxyestradiol (2-ME) reduces atherosclerotic lesion formation, while the underlying mechanisms remain obscure. In this work, we investigated the vasodilatory effect of 2-ME and the role of nitric oxide (NO) involved. In vivo studies using noninvasive tail-cuff methods showed that 2-ME decreased blood pressure in Sprague Dawley rats. Furthermore, in vitro studies showed that cumulative addition of 2-ME to the aorta caused a dose- and endothelium-dependent vasodilation. This effect was unaffected by the pretreatment with the pure estrogen receptor antagonist ICI 182,780, but was largely impaired by endothelial nitric oxide synthase (eNOS) inhibitor NG-nitro-L-arginine methyl ester (L-NAME) or by phosphoinositide 3-kinase (PI3K) inhibitor wortmannin (WM). Moreover, 2-ME（10-7 ∼10-5 M）enhanced phosphorylation of Akt and eNOS and promoted NO release from cultured human umbilical endothelial cells (HUVECs). These effects were blocked by PI3K inhibitor WM, or by the transfection with Akt specific siRNA, indicating that endothelial Akt/eNOS/NO cascade plays a crucial role in 2-ME-induced vasodilation. The peroxisome proliferator-activated receptor γ (PPARγ) mRNA and protein expression were detected in HUVECs and the antagonist GW9662 or the transfection with specific PPARγ siRNA inhibited 2-ME-induced eNOS and Akt phosphorylation, leading to the impairment of NO production and vasodilation. In conclusion, 2-ME induces vasodilation by stimulating NO release. These actions may be mediated by PPARγ and the subsequent activation of Akt/eNOS cascade in vascular endothelial cells.

KGFR promotes Na+ channel expression in a rat acute lung injury model.

BACKGROUND: Binding of keratinocyte growth factor (KGF) to the KGF receptor (KGFR) plays an important role in the recovery of alveolar epithelial cells from acute lung injury (ALI). OBJECTIVES: To evaluate the effect of gene therapy via adenovirus gene transfer of KGFR on the treatment of ALI. METHODS: Sprague-Dawley rats were divided into four groups: normal controls, injury controls, normal adenovirus transduced group and injury adenovirus transduced group. The ALI model was induced by lipopolysaccharide (LPS) injection. Recombinant adenovirus (AdEasy-KGFR) was injected via the tail vein. Expression of the sodium (Na(+)) channel in rat alveolar type II (ATII) epithelial cells was determined by PCR, immunohistochemistry and immunoelectron microscopy of rat lung tissues. RESULTS: Gene expression of the Na(+) channel and KGFR in ATII cells was higher in the normal adenovirus transduced group than the three other groups; expression of these two genes in the injury adenovirus transduced group was higher than the injury control group. Na(+) channel protein expression was lower in the injury adenovirus transduced group but higher than the injury control group. CONCLUSIONS: KGFR over-expression induced Na channel expression could potentially be beneficial for ALI therapy.

17ß-estradiol induces vasorelaxation by stimulating endothelial hydrogen sulfide release.

Estrogen exerts vascular protective effects, but the underlying mechanisms remain to be understood fully. In recent years, hydrogen sulfide (H(2)S) has increasingly been recognized as an important signaling molecule in the cardiovascular system. Vascular H(2)S is produced from L-cysteine, catalyzed by cystathionine γ-lyase (CSE). In our study, apolipoprotein E (ApoE)-deficient mice were ovariectomized and implanted with placebo (OVX mice) or 17ß-estradiol (E(2)) pellets (OVX + E(2) mice). Compared with OVX mice, OVX + E(2) mice showed increased plasma H(2)S levels (P = 0.012) and decreased aortic lesion area (P = 0.028). These effects were largely reversed when supplementing with the irreversible CSE inhibitor DL-propargylglycine (PPG) in the OVX + E(2) + PPG mice. Meanwhile, the nitric oxide and prostacyclin-resistant responses to cumulative application of acetylcholine (ACh) were studied among all the three groups of femoral arteries. Compared with the arteries in the OVX group, the vasodilator sensitivity of arteries to ACh was increased in the OVX + E(2) group and attenuated in the OVX + E(2) + PPG group. E(2) and estrogen receptor (ER) α agonist 4',4″,4'″-(4-propyl-[1H]-pyrazole-1,3,5-triyl) trisphenol rapidly increased H(2)S release in human endothelial cells, but not partially selective ERß agonist 2,3-bis-(4-hydroxyphenyl)-propionitrile. These effects were inhibited by ER antagonist ICI 182780 or by protein kinase G (PKG) inhibitor KT5823. Furthermore, endothelial PKG activity was increased by E(2) (P = 0.003) and E(2)-induced vasodilation was inhibited by KT5823 (P = 0.009). In conclusion, the endothelial CSE/H(2)S pathway is activated by E(2) through PKG, which leads to vasodilation. These actions may be relevant to estrogen's anti-atherogenic effect.