Polysaccharides from waste Zingiber mioga leaves: Ultrasonic-microwave-assisted extraction, characterization, antioxidant and anticoagulant potentials.

Zingiber mioga is a highly economic crop that is used to produce vegetables, spices and herbal pharmaceuticals. Its edible flower bud contributes most to the economic value, but the big leaves were discarded as agricultural waste, which urgently needs to be exploited. In this work, polysaccharides from waste Z. mioga leaves (PWZMLs) were extracted using ultrasonic-microwave-assisted extraction (UMAE). After purification and characterization, the antioxidation and anticoagulation of PWZMLs were evaluated to appraise the potential in cardiovascular protection. Under the liquid-solid ratio of 26: 1 mL/g, after ultrasonication at 495 W for 10 min, followed by microwaving at 490 W for 5 min, the yield of PWZMLs achieved to 6.22 ± 0.14 %, notably higher (P < 0.01) than other methods, and ultrasound contributed more to the yield than microwave. Various analyses confirmed that PWZMLs were negatively charged polysaccharides with galacturonic acid the dominant uronic acid. PWZMLs exerted excellent antioxidant capacity, especially for scavenging 1, 1-diphenyl-2-picrylhydrazyl radical. PWZMLs also elicited promising anticoagulant property, particularly for prolonging activated partial thromboplastin time and lowering fibrinogen, which were almost equivalent to heparin at the same concentration. PWZMLs contained two polysaccharide fractions (199.53 and 275.42 kDa) that could synergistically contribute to the pronounced antioxidant and anticoagulant activities. The PWZMLs extracted with optimized UMAE have great potential in cardiovascular protection.

The Immunosuppressive Roles of PD-L1 during Influenza A Virus Infection.

The clinical benefits of targeting programmed death-ligand 1 (PD-L1) in various cancers represent a strategy for the treatment of immunosuppressive diseases. Here, it was demonstrated that the expression levels of PD-L1 in cells were greatly upregulated in response to H1N1 influenza A virus (IAV) infection. Overexpression of PD-L1 promoted viral replication and downregulated type-I and type-III interferons and interferon-stimulated genes. Moreover, the association between PD-L1 and Src homology region-2, containing protein tyrosine phosphatase (SHP2), during IAV/H1N1 infection was analyzed by employing the SHP2 inhibitor (SHP099), siSHP2, and pNL-SHP2. The results showed that the expressions of PD-L1 mRNA and protein were decreased under SHP099 or siSHP2 treatment, whereas the cells overexpressing SHP2 exhibited the opposite effects. Additionally, the effects of PD-L1 on the expression of p-ERK and p-SHP2 were investigated in PD-L1-overexpressed cells following WSN or PR8 infection, determining that the PD-L1 overexpression led to the decreased expression of p-SHP2 and p-ERK induced by WSN or PR8 infection. Taken together, these data reveal that PD-L1 could play an important role in immunosuppression during IAV/H1N1 infection; thus, it may serve as a promising therapeutic target for development of novel anti-IAV drugs.

5-ALA-PDT induced ferroptosis in keloid fibroblasts via ROS, accompanied by downregulation of xCT, GPX4.

Keloids display many cancerous properties, including uncontrolled and invasive growth, high rates of recurrence as well as similar bioenergetics. 5-aminolevulinic acid-based photodynamic therapy (5-ALA-PDT) is an effective treatment that performs cytotoxic effects by producing reactive oxygen species (ROS), which is linked to lipid peroxidation and ferroptosis. Herein, we explored underlying mechanisms of 5-ALA-PDT against keloids. We identified that 5-ALA-PDT led to elevated levels of ROS and lipid peroxidation in keloid fibroblasts, accompanied by downregulation of xCT and GPX4, which are associated with anti-oxidation effects and ferroptosis inhibition. These results may indicate that 5-ALA-PDT treatment increases ROS while inhibiting xCT and GPX4, thus promoting lipid peroxidation to induce ferroptosis in keloid fibroblasts.

First-in-human phase I/Ib study of QL1706 (PSB205), a bifunctional PD1/CTLA4 dual blocker, in patients with advanced solid tumors.

BACKGROUND: QL1706 (PSB205) is a single bifunctional MabPair (a novel technical platform) product consisting of two engineered monoclonal antibodies (anti-PD-1 IgG4 and anti-CTLA-4 IgG1), with a shorter elimination half-life (t1/2) for CTLA-4. We report results from a phase I/Ib study of QL1706 in patients with advanced solid tumors who failed standard therapies. METHODS: In the phase I study, QL1706 was administered intravenously once every 3 weeks at one of five doses ranging from 0.3 to 10 mg/kg, and the maximum tolerated dose, recommended phase 2 dose (RP2D), safety, pharmacokinetics (PK), and pharmacodynamics (PD) of QL1706 were investigated. In the phase Ib study, QL1706 was administered at the RP2D intravenously every 3 weeks, and the preliminary efficacies in non-small cell lung cancer (NSCLC), nasopharyngeal carcinoma (NPC), cervical cancer (CC), and other solid tumors were evaluated. RESULTS: Between March 2020 and July 2021, 518 patients with advanced solid tumors were enrolled (phase I, n = 99; phase Ib, n = 419). For all patients, the three most common treatment-related adverse events (TRAEs) were rash (19.7%), hypothyroidism (13.5%), and pruritus (13.3%). The TRAEs and immune-related adverse events (irAEs) of grade ≥ 3 occurred in 16.0% and 8.1% of patients, respectively. In phase I, 2 of 6 patients in the 10mg/kg group experienced dose-limiting toxicities (DLTs) (grade 3 thrombocytopenia and grade 4 immune-mediated nephritis), so the maximum tolerated dose (MTD) was reached at 10 mg/kg. The RP2D was determined to be 5 mg/kg based on comprehensive analysis of tolerability, PK/PD, and efficacy. For all patients who received QL1706 at the RP2D, the objective response rate (ORR) and median duration of response were 16.9% (79/468) and 11.7 months (8.3-not reached [NR]), respectively; and the ORRs were 14.0% (17/121) in NSCLC, 24.5% (27/110) in NPC, 27.3% (15/55) in CC, 7.4% (2/27) in colorectal cancer, 23.1% (6/26) in small cell lung cancer. For immunotherapy-naive patients, QL1706 exhibited promising antitumor activities, especially in NSCLC, NPC, and CC, with ORRs of 24.2%, 38.7%, and 28.3%, respectively. CONCLUSIONS: QL1706 was well tolerated and demonstrated promising antitumor activity in solid tumors, especially in NSCLC, NPC, and CC patients. It is currently being evaluated in randomized phase II (NCT05576272, NCT05179317) and phase III (NCT05446883, NCT05487391) trials. Trial Registration ClinicalTrials.gov Identifier: NCT04296994 and NCT05171790.

RGC-32 facilitates pancreatic cancer via activating Wnt/ß-catenin signaling.

Longitudinal studies have indicated the facilitating effect of RGC-32 during diverse disease progression including pancreatic cancer, yet the systematic and detailed effect of RGC-32 during pancreatic cancer is largely unknowable. For this purpose, we took advantage of the pancreatic cancer cell line (BXPC3) with RGC-32 expression and then modulated its expression by lentivirus-mediated knockdown (shRGC-32) and overexpression (pcDNA-RGC-32). To verify the effect of Wnt/ß-catenin signaling in RGC-32-based tumorigenicity, we added the agonist CT99021 to the shRGC-32 BXPC3 cell line and pancreatic cancer mouse model. The deficiency in cellular vitality (cell survival, apoptosis, cell cycle) and migration of BXPC3 were sharply rescued by shRGC-32 in vitro. Notably, the aforementioned phenotypes as well as the expression pattern of EMT-associated biomarkers of BXPC3 with shRGC-32 expression could largely rescued by the agonist of Wnt/ß-catenin in vitro and in vivo. Our data indicated the facilitating effect of RGC-32 upon pancreatic cancer cell line and mouse model via activating the Wnt/ß-catenin signaling, which collectively suggested the feasibility of RGC-32 as a potent diagnostic and therapeutic target of pancreatic cancer in the future.

Cysteine Donor-Based Brain-Targeting Prodrug: Opportunities and Challenges.

Overcoming blood-brain barrier (BBB) to improve brain bioavailability of therapeutic drug remains an ongoing concern. Prodrug is one of the most reliable approaches for delivering agents with low-level BBB permeability into the brain. The well-known antioxidant capacities of cysteine (Cys) and its vital role in glutathione (GSH) synthesis indicate that Cys-based prodrug could potentiate therapeutic drugs against oxidative stress-related neurodegenerative disorders. Moreover, prodrug with Cys moiety could be recognized by the excitatory amino acid transporter 3 (EAAT3) that is highly expressed at the BBB and transports drug into the brain. In this review, we summarized the strategies of crossing BBB, properties of EAAT3 and its natural substrates, Cys and its donors, and Cys donor-based brain-targeting prodrugs by referring to recent investigations. Moreover, the challenges that we are faced with and future research orientations were also addressed and proposed. It is hoped that present review will provide evidence for the pursuit of novel Cys donor-based brain-targeting prodrug.

Neuropeptide S and its receptor NPSR enhance the susceptibility of hosts to pseudorabies virus infection.

The neuropeptide S (NPS) and its receptor (NPSR) represent a signaling system in the brain. Increased levels of NPS and NPSR have been observed in PK15 cells and murine brains in response to pseudorabies virus (PRV) infection, but it remains unclear whether elevated levels of NPS and NPSR are involved in the pathogenic process of PRV infection. In this study, the activities of both NPS and NPSR during PRV pathogenesis were explored in vitro and in vivo by reverse transcription polymerase chain reaction (RT-PCR), PCR, real-time quantitative RT-PCR (qRT-PCR), qPCR, TCID50, and Western blotting methods. NPSR-deficient cells were less susceptible to PRV infection, as evidenced by decreased viral production and PRV-glycoprotein E (gE) expression. In vitro studies showed that exogenous NPS promoted the expression of interleukin 6 (IL-6) mRNA but inhibited interferon ß (IFN-ß) mRNA expression in PK15 cells after PRV infection. In vivo studies showed that NPS-treated mice were highly susceptible to PRV infection, with decreased survival rates and body weights. In addition, NPS-treated mice showed elevated levels of IL-6 mRNA and STAT3 phosphorylation. However, the expression of IFN-ß mRNA was greatly decreased after virus challenge. Contrasting results were obtained from the NPSR-ir-treated groups, which further highlighted the effects of NPS. This study revealed that NPS-treated hosts are more susceptible to PRV infection than controls. Moreover, excessive IL-6/STAT3 and defective IFN-ß responses in NPS-treated mice may contribute to the pathogenesis of PRV.

Eleocharis dulcis corm: phytochemicals, health benefits, processing and food products.

Eleocharis dulcis, an aquatic plant belonging to Cyperaceae family, is indigenous to Asia, and also occurs in tropical Africa and Australia. The edible corm part of E. dulcis is a commonly consumed aquatic vegetable with a planting area of 44.46 × 103 hm2 in China. This work aims to explore the potential of E. dulcis corm for use as a new food source for sufficient nutrients and health benefits by reviewing its nutrients, phytochemicals, functions, processing and food products. Eleocharis dulcis corm contains starches, dietary fibers, non-starch polysaccharides, proteins, amino acids, phenolics, sterols, puchiin, saponins, minerals and vitamins. Among them, phenolics including flavonoids and quinones could be the major bioconstituents that largely contribute to antioxidant, anti-inflammatory, antibacterial, antitumor, hepatoprotective, neuroprotective and hypolipidemic functions. Peel wastes of E. dulcis corm tend to be enriched in phenolics to a much higher extent than the edible pulp. Fresh-cut E. dulcis corm can be consumed as a ready-to-eat food or processed into juice for beverage production, and anti-browning processing is a key to prolonging shelf life. Present food products of E. dulcis corm are centered on various fruit and vegetable beverages, and suffer from single categories and inadequate development. In brief, underutilized E. dulcis corm possesses great potential for use as a new food source for sufficient nutrients and health benefits. © 2021 Society of Chemical Industry.

A modified urinary catheterisation technique to reduce urine leakage: A prospective randomised study.

BACKGROUND: Long-term indwelling catheters assist people who are unable to use another bladder management method. However, urine leakage is a common problem with an indwelling urinary catheter. This study aims to determine whether a modified catheterisation technique would reduce urine leakage incidence. METHODS: Participants were randomly divided into conventional or modified catheterisation groups. In the modified technique group, the volume of fluid that needed to be injected into the balloon to obtain a suitable catheter front-end curvature (120-145°) was measured before catheterisation. Baseline characteristics and first-time success rates and procedure durations were similar between groups. RESULTS: There were 30 patients in each group. Compared with conventional catheterisation, the modified catheterisation group had smaller residual urine volume (median 11 mL Vs. 30.5 mL, p<0.001) and more leakage-free days (30 days Vs. 10 days, p<0.001). Leakage-free survival was longer in the modified catheterisation group (p<0.001). The residual urine volume (>17 vs ≤17 ml (median); incident rate ratio (IRR), 28.710; 95%CI, 4.114-200.331; p=0.001) was independently associated with urine leakage. CONCLUSIONS: The modified catheterisation technique may reduce the incidence of urine leakage.

The consistency of an optical body surface scanning method compared with computed tomography: a validation study.

OBJECTIVE: The ideal noninvasive method for evaluation of pectus excavatum remains to be defined. We sought to verify the accuracy of an optical body surface scanning method compared with conventional CT scan. MATERIALS AND METHODS: A PrimeSense 3D sensor was used to obtain data from patients undergoing surgical or noninvasive treatment for pectus excavatum. The Haller index, external Haller index, and depth ratio were then calculated from both body scan and computed tomography scan data for the same patients. Statistical analyses were carried out to find if there is consistency between data from body scanning and computed tomography. RESULTS: Data acquisition was complete. In total, 40 patients (median age: 5.03 years, 11 female) with pectus excavatum undergoing nonoperative (n = 13) or surgical Nuss treatment (n = 27) were included. The Haller index was lower in vacuum bell patients, which also had a higher female proportion. Pearson correlation coefficient between external Haller indices from body scanning and from computed tomography and between the depth ratios from body scanning and from computed tomography were 0.63 and 0.84, respectively. By intraclass correlation coefficient method, the correlation coefficient was 0.56 between external Haller indices from body scanning and from computed tomography and 0.80 between depth ratios from body scanning and from computed tomography. CONCLUSION: The optical body surface scanning is a reliable approach to the measurement of PE severity and could be routinely used in the monitoring of PE development of treatment, especially in the pediatric population. STUDY TYPE: Diagnostic test. LEVEL OF EVIDENCE: Level II.

EphB4 inhibitor overcome the acquired resistance to cisplatin in melanomas xenograft model.

The purpose of this paper is to investigate the possible mechanisms of resistance to chemotherapy in melanoma from the perspective of molecular biology and to discuss the strategies to overcome them. Cisplatin, a DNA-damaging compound that triggers apoptotic cell death, is commonly used in the treatment of malignant melanoma. However, most patients develop mechanisms of acquired resistance and about 25% of them do not achieve tumor regression at all, due to intrinsic resistance to therapy. In the current study, we reported the tumor xenografts of the human A375 melanoma, after 40-weeks' consecutive therapy with cisplatin that developed resistance as a result of EphB4 overexpression. Moreover, the expression of phospho-AKT and phospho-ERK were significantly increased in cisplatin-resistant tumors. In addition, combined of cisplatin with EphB4 selective inhibitor could abrogate this acquired mechanism of drug resistance due to an enhanced apoptotic effect in cisplatin-resistant xenografts. In summary, these results help to understand the mechanisms of acquired resistance to chemotherapy and provide important information for clinical treatment strategies.

SCF/c-kit signaling is required in 12-O-tetradecanoylphorbol-13-acetate-induced migration and differentiation of hair follicle melanocytes for epidermal pigmentation.

Hair follicle melanocyte stem cells (McSCs) are responsible for hair pigmentation and also function as a major melanocyte reservoir for epidermal pigmentation. However, the molecular mechanism promoting McSCs for epidermal pigmentation remains elusive. 12-O-tetradecanoylphorbol-13-acetate (TPA) mimics key signaling involved in melanocyte growth, migration and differentiation. We therefore investigated the molecular basis for the contribution of hair follicle McSCs to epidermal pigmentation using the TPA induction model. We found that repetitive TPA treatment of female C57BL/6 mouse dorsal skin induced epidermal pigmentation by increasing the number of epidermal melanocytes. Particularly, TPA treatment induced McSCs to initiate proliferation, exit the stem cell niche and differentiate. We also demonstrated that TPA promotes melanoblast migration and differentiation in vitro. At the molecular level, TPA treatment induced robust expression of stem cell factor (SCF) in keratinocytes and c-kit in melanoblasts and melanocytes. Administration of ACK2, a neutralizing antibody against the Kit receptor, suppressed mouse epidermal pigmentation, decreased the number of epidermal melanocytes, and inhibited melanoblast migration. Taken together, our data demonstrate that TPA promotes the expansion, migration and differentiation of hair follicle McSCs for mouse epidermal pigmentation. SCF/c-kit signaling was required for TPA-induced migration and differentiation of hair follicle melanocytes. Our findings may provide an excellent model to investigate the signaling mechanisms regulating epidermal pigmentation from mouse hair follicle McSCs, and a potential therapeutic option for skin pigmentation disorders.

Anti-tumor effect of polysaccharides from Scutellaria barbata D. Don on the 95-D xenograft model via inhibition of the C-met pathway.

Polysaccharides isolated from Scutellaria barbata (PSB) have been reported to have anti-tumor effects. To investigate the underlying mechanism, a highly invasive, metastatic and phospho-c-Met overexpression lung carcinoma cell, 95-D cell line was used. The results showed that in vitro, PSB not only could inhibit the proliferation of 95-D cell line (IC(50) = 35.2 µg/mL), but also down-regulated the expression of phospho-c-Met and its downstream signaling molecules including phospho-Erk and phospho-Akt. In vivo, PSB inhibited tumor growth in the 95-D subcutaneous xenograft model in a dose-dependent manner; after once-daily intraperitoneal injection for 3 weeks, tumor growth inhibition T/C ratio for 100 and 200 mg/kg treatments was 42.72% and 13.6%, respectively. In the end of the in vivo study, tumor tissues were harvested for further evaluation of the phosphorylation level of c-Met, AKT, and ERK. Ex vivo results demonstrated that the phosphorylation of c-Met and its downstream signaling molecules were also significantly inhibited by PSB. Immunohistochemistry analysis showed dose-dependent inhibition of tumor cell proliferation (Ki67) and reduction of microvessel density (CD31). In summary, the results indicated that PSB exerted anti-tumor growth activity on human lung cancer 95-D in vitro and in vivo by directly regulating the c-Met signaling pathway and the anti-tumor effects were mainly based on its anti-proliferation and anti-angiogenesis action.

Research on tumorigenicity of cinnamaldehyde in melanoma cell lines and its mechanism.

Melanoma is a highly malignant tumor originating from melanocytes. This disease is characterized by inconspicuous onset, high malignancy, and poor prognosis. The aim of this study is to explore the effect of cinnamaldehyde on melanoma tumorigenicity and its mechanism. Melanoma cells were subcutaneously injected into a nude mouse to establish the tumour model. A comparison was made for the difference in formation and growth of melanoma cell tumor between normal saline and cinnamaldehyde. A comparison was also made for the number of new vessels between the normal saline group (the control group) and the cinnamaldehyde group (the experimental group) through immumohistochemical staining. The western blot was used to detect the difference in expression levels of vascularization related proteins. The results indicated that the volume of tumors formed and the number of new vessels in melanoma cells of the cinnamaldehyde group decreased significantly compared with those in the cells of the normal saline group. A further study indicated that the expression of hypoxia-inducible factor-a (HIF-α) and vascular endothelial growth factor (VEGF) in the melanoma of the cinnamaldehyde group decreased significantly. In conclusion, cinnamaldehyde plays a certain role in inhibiting the occurrence and progression of melanoma and its action mechanism may be manifested by inhibiting expression of VEGF and HIF-α, thus blood vessel simulation and formation of new blood vessels of melanoma cells, and growth of tumors accordingly.

Quantitative assessment of the association between XPC Lys939Gln polymorphism and cutaneous melanoma risk.

Previous studies evaluating the association between XPC Lys939Gln polymorphism and cutaneous melanoma risk reported conflicting findings. We searched PubMed and Embase databases up to May 16, 2013 to identify eligible studies on the association between XPC Lys939Gln polymorphism and cutaneous melanoma risk. Finally, a total of seven case-control studies including 3,971 cases of cutaneous melanoma and 5,873 controls were included in the meta-analysis. Statistical analysis was performed with STATA version 11.0. Odds ratios (ORs) with 95 % confidence intervals (95 % CIs) were used to assess the strength of the association. Overall, there was no association between XPC Lys939Gln polymorphism and cutaneous melanoma risk under all five genetic models (Gln vs. Lys: OR = 1.11, 95 % CI = 0.98-1.26, P = 0.10; GlnGln vs. LysLys: OR = 1.26, 95 % CI = 0.98-1.61, P = 0.07; LysGln vs. LysLys: OR = 1.04, 95 % CI = 0.88-1.22, P = 0.64; GlnGln/LysGln vs. LysLys: OR = 1.10, 95 % CI = 0.92-1.31, P = 0.29; GlnGln vs. LysLys/LysGln: OR = 1.19, 95 % CI = 0.99-1.43, P = 0.06). Subgroup analysis in Caucasians showed that there was an obvious association between XPC Lys939Gln polymorphism and cutaneous melanoma risk in Caucasians (GlnGln vs. LysLys/LysGln: OR = 1.12, 95 % CI = 1.00-1.25, P = 0.05). Sensitivity analysis by omitting one study in turns showed that the significance of the pooled ORs was not stable. In addition, there was some evidence of publication bias in the meta-analysis, and meta-analyses of the studies with large sample size did not find the obvious association between XPC Lys939Gln polymorphism and cutaneous melanoma risk in Caucasians. Therefore, there is little evidence for the association between XPC Lys939Gln polymorphism and cutaneous melanoma risk.

[Experimental study on constructing a tissue engineered epidermis for replacement of skin irritation in vitro].

OBJECTIVE: To investigate the method of constructing a tissue engineered epidermis with human epidermal cells and polycarbonate membrane, and to establish a tissue engineered epidermis with barrier function which is intended to be the replacing model in vitro of skin irritation test. METHODS: The tissue engineered epidermis was constructed by using polycarbonate membrane as scaffold and stratified differentiated epidermis derived from human keratinocytes. The tissue engineered epidermis was cultured on an inert polycarbonate filter at the air-liquid interface. After 13 days of culture, the composition and structure of tissue engineered epidermis were observed by HE staining, immunofluorescence staining of keratin 10 (K10) & K13, K14, laminin, involucrin, and filaggrin, and transmission electronic microscope. The half maximal inhibitory concentration of a substance (IC50) of SDS was determined in the penetration test of tissue engineered epidermis cultured in the absence (control group) or the presence (experimental group) of lipid supplement for 18 hours. RESULTS: The constructed epidermis was similar to normal epidermis, which was consisted of a proliferating basal layer, differentiated spinous layer, granular layer, and stratum corneum. The IC50 values of tissue engineered epidermis cultured in the control group and experimental group were 0.072% (2.36 mmol/L) and 0.183% (6.00 mmol/L), respectively. CONCLUSION: The tissue engineered epidermis constructed on polycarbonate membrane has normal composition and structure and barrier function corresponding to the normal epidermis.

Effects of neuropeptide S on the proliferation of splenic lymphocytes, phagocytosis, and proinflammatory cytokine production of pulmonary alveolar macrophages in the pig.

Neuropeptide S (NPS), a newly identified neuropeptide, is involved in many physiological and pathological activities through the NPS receptor (NPSR). Recently, the NPS and NPSR have been detected in peripheral systems of pigs including immune tissues, suggesting that NPS may play an important role in the regulation of immune function. The aim of this study was to demonstrate the presence and function of NPS and NPSR in splenic lymphocytes (SPLs) and pulmonary alveolar macrophages (PAMs) of pigs. By RT-PCR, the expression of NPS and NPSR mRNA was detected in the SPLs and PAMs. NPS immunoreactivity was observed in the membrane and cytoplasm of both SPLs and PAMs. We found that NPS could stimulate the proliferation of SPLs, when NPS was added at concentrations of 0.01, 0.1, 1, 10, 100 and 1000 nM alone or in combination with PHA/LPS in vitro. In macrophages from bronchoalveolar lavage (BAL) fluid of pigs, various doses of NPS (0.01, 0.1, 1, 10, 100 and 1000 nM) up-regulated the phagocytosis of PAMs in comparison to controls. In PAMs, NPS could induce the production of the pro-inflammatory cytokines IL-1ß, IL-6 and TNF-α. Taken together, all data suggest that NPS is capable of inducing phagocytosis of non-opsonized E. coli. NPS might act as potent neuroimmunomodulatory factors and affects the maintenance of immune homeostasis.

The regulatory mechanism of neuromedin S on luteinizing hormone in pigs.

Neuromedin S (NMS) has been implicated in the regulation of luteinizing hormone (LH) secretion. However, the regulatory mechanism of NMS on LH in pigs remains unexplored. In the present study, we confirmed the hypothesis that the effect of NMS on LH could be mediated via hypothalamic melanocyte-stimulating hormones (MSH) neurons of ovariectomized pigs. In an immunohistological experiment, NMS receptor NMU2R-positive neurons were found in the paraventricular nucleus of hypothalamus, widely distributed in the anterior pituitary, and sparsely observed in the posterior pituitary. We also found that serum LH level was declined at between 12 and 60 min with the lowest level at 24 min after NMS injection. The decreased LH secretion induced by NMS could be completely abolished by pretreatment with melanocortin receptor-4 antagonist SHU9119, while a signal injection of 1.0 nM SHU9119 per se did not affect the serum LH level. Real time quantitative RT-PCR results showed that the expression of GnRH and LH mRNAs were down-regulated by NMS treatment, but their reduction was restored to normal level by SHU9119 treatments. The expression of NMU2R and PR mRNAs were up-regulated by NMS treatment, but their effects were blocked by SHU9119 treatments. The expression of the estrogen receptor mRNA in the pig hypothalamus and pituitary was unchanged under the NMS and SHU9119+NMS treatments. In summary, all results suggest that the inhibitory effect of NMS on LH is at least in part through its receptor NMU2R and mediated via MSH neurons in hypothalamus-pituitary axis of ovariectomized pigs.

Expression of NMS and NMU2R in the pig reproductive axis during the estrus cycle and the effect of NMS on the reproductive axis in vitro.

Evidence has revealed that neuromedin S (NMS) and neuromedin U-receptor type-2 (NMU2R) mRNAs are expressed in the central nervous system and reproductive organs. Previous data indicated that variation of NMS and NMU2R was due to the phases of the adult rat hypothalamus estrus cycle. However, the expression and function of NMS in the pig reproductive axis remains unexplored. In this study, 16 virginal gilts were classified into four groups: proestrus, estrus, diestrus 1, and diestrus 2; the expression of NMS and NMU2R in the cyclic pig hypothalamus-pituitary-ovary axis was studied by reverse transcriptaion-polymerase chain reaction (RT-PCR), and the effect of NMS on the reproductive axis in vitro was detected by radioimmunoassay (RIA). The cloned pig NMS and NMU2R sequences were 82% and 90.2% identical to those of the corresponding human homologues, respectively. RT-PCR showed that NMS and NMU2R mRNA expression in the hypothalamus and pituitary changed with the estrus cycle, i.e., with the highest level in the proestrus group and the lowest in the estrus group. In the ovary, NMS and NMU2R expression was highest in the diestrus 2 group and the lowest in the proestrus group. In the in vitro study, different concentrations of NMS induced the release of gonadotropin-releasing hormone, luteinizing hormone, and estradiol at different levels of the reproductive axis. Taken together, the expression pattern of NMS during the estrus cycle and its role in reproductive hormones in vitro provide novel evidences of the potential roles of NMS in the regulation of pig reproduction.

[Influence of Wnt-1 recombinant adenovirus on differentiation of human epidermal stem cells].

OBJECTIVE: To observe the influence of Wnt-1 recombinant adenovirus on differentiation tendency of human epidermal stem cells. METHODS: Wnt-1 recombinant adenovirus was transduced into hESCs (E group), while normal hESCs were used as control (C) group. The diameter, proliferation,and labeling molecular expression of hESC were determined. The content of MMP-2 and MMP-7 in supernate were also assayed. RESULTS: There was no obvious difference in diameter of hESC between two groups. The density of hESC in E group was (1.45 +/- 0.09) x 10(5)/mL, which was obviously higher than that in C group [(1.18 +/- 0.10) x 10(5)/mL, P < 0.05]. There were no obvious differences in expression of markers between two groups,including keratin 5 (KS), K6, K7, KS, K14, CD44, carcinoembryonic-like antigen (CEAA), ER, PR (P > 0.05) ,while the expression of K 10 was different among groups [(60 +/- 3)% in E group, 0 in C group], also K18 [(34.3 +/- 2.1)% in E group vs. (13.8 +/- 1.7)% in C group, P < 0.05], and K19 [(17.1 +/- 1.8)% in E group vs. (24.4 +/- 1.5)% in C group, P < 0.05].The contents of MMP-2 and MMP-7 in E group were higher than those in C group (P < 0.01). CONCLUSION: Wnt-1 recombinant adenovirus can induce the differentiation of hESCs to glandular epithelium-like cells.