Overexpression of LVRN impedes the invasion of trophoblasts by inhibiting epithelial-mesenchymal transition.

Laeverin (LVRN) was first detected on the outer layer of the chorion laeve and migrating extravillous trophoblasts (EVTs). It is an enzyme that plays an important role in the placentation and pathophysiology of preeclampsia (PE). Previous studies have indicated that LVRN may be required for the invasion of human trophoblast cells. Paradoxically, LVRN was found to be highly expressed in the trophoblasts of PE patients with impaired invasive capacities. In this study, we detected the expression of LVRN in the placentas of PE patients (n=5) and normal term pregnancy women (n=5) as a control group by immunohistochemistry. LVRN was elevated in decidua (P=0.0083) and villi (P=0.0079) of PE patients. Next, LVRN was overexpressed via adeno-associated virus-mediated gene transfer in trophoblastic cell lines HTR8, Swan71, and JAR. Matrigel transwell assay and wound healing assay showed that overexpression of LVRN impeded the invasion of these three cell lines. Western blot analysis showed that LVRN overexpression caused downregulation of N-cadherin and vimentin and upregulation of E-cadherin, suggesting the inhibitory role of LVRN in epithelial-mesenchymal transition (EMT). Moreover, our data indicated that long noncoding RNA NONSTAT103348 (lnc10-7) was elevated in PE patients. Silencing lnc10-7 led to decreased LVRN expression. Taken together, although the basal level of LVRN may be crucial for cell invasion, overexpression of LVRN may abrogate the cell invasiveness, suggesting a multifaceted role of LVRN in the pathogenesis of PE.

Increased leptin-b expression and metalloprotease expression contributed to the pyridoxine-associated toxicity in zebrafish larvae displaying seizure-like behavior.

Epilepsy is a common neurological disorder affecting people of all ages, races and ethnic backgrounds world-wide. Vitamin B6 supplementation has been widely used as an adjuvant for treating epilepsy. However, the adverse effects, including nausea and peripheral sensory neuropathy, caused by long-term and high-dose consumption of vitamin B6 have undermined the usefulness of vitamin B6 supplementation, justifying additional experimental scrutiny of vitamin B6-associated toxicity. In the current study, we found that the presence of pyridoxine, the inactive form of B6 vitamer included in most nutrient supplements, increased the mortality of the larvae displaying chemical-induced epilepsy. The expression of leptin-b, one zebrafish ortholog of human leptin, was significantly increased in the larvae displaying seizures. Increased leptin-b expression alleviated larval seizure-like behavior when exposed to epilepsy inducer, but also increased larval mortality in the presence of pyridoxine. Meanwhile, elevated adam17 and mmp13 mRNA level were found in the larvae simultaneously exposed to epilepsy-inducer and pyridoxine. Adding TNF-α inhibitor and mmp13 inhibitor effectively improved the survival of larvae injected with leptin-b mRNA and exposed to pyridoxine subsequently. We conclude that increased leptin-b and metalloprotease expression contributed, at least partly, to the pyridoxine-associated toxicity observed in larvae displaying seizures.

Bioinformatic characterization of angiotensin-converting enzyme 2, the entry receptor for SARS-CoV-2.

The World Health Organization declared the COVID-19 epidemic a public health emergency of international concern on March 11th, 2020, and the pandemic is rapidly spreading worldwide. COVID-19 is caused by a novel coronavirus SARS-CoV-2, which enters human target cells via angiotensin converting enzyme 2 (ACE2). We used a number of bioinformatics tools to computationally characterize ACE2 by determining its cell-specific expression in trachea, lung, and small intestine, derive its putative functions, and predict transcriptional regulation. The small intestine expressed higher levels of ACE2 mRNA than any other organ. By immunohistochemistry, duodenum, kidney and testis showed strong signals, whereas the signal was weak in the respiratory tract. Single cell RNA-Seq data from trachea indicated positive signals along the respiratory tract in key protective cell types including club, goblet, proliferating, and ciliary epithelial cells; while in lung the ratio of ACE2-expressing cells was low in all cell types (<2.6%), but was highest in vascular endothelial and goblet cells. Gene ontology analysis suggested that, besides its classical role in the renin-angiotensin system, ACE2 may be functionally associated with angiogenesis/blood vessel morphogenesis. Using a novel tool for the prediction of transcription factor binding sites we identified several putative binding sites within two tissue-specific promoters of the ACE2 gene as well as a new putative short form of ACE2. These include several interferon-stimulated response elements sites for STAT1, IRF8, and IRF9. Our results also confirmed that age and gender play no significant role in the regulation of ACE2 mRNA expression in the lung.

Comparative analysis of the expression of metalloproteases (MMP-2, MMP-9, MMP-11 and MMP-13) and the tissue inhibitor of metalloprotease 3 (TIMP-3) between previous negative biopsies and radical prostatectomies. / Análisis comparativo de la expresión de metaloproteasas (MMP-2, MMP-9, MMP-11 y MMP-13) y del inhibidor tisular de la metaloproteasa 3 (TIMP-3) entre las biopsias previas negativas y las prostatectomías radicales.

Metalloproteases (MMPs) and tissue inhibitor of metalloprotease-3 (TIMP-3) have been associated to the risk of having cancer and tumor aggressiveness. When facing the difficulties of prostate cancer diagnosis, the expression of MMPs and TIMP-3 in negative biopsies could be helpful to evaluate a diagnostic suspicion. Our objective is to carry out a comparative study of the expression of MMPs and TIMP-3 in previous negative biopsies and radical prostatectomies (RP). MATERIAL AND METHODS: Retrospective analysis of a hospital-based cohort including 21 patients with suspicion of prostate carcinoma, whose expressions of MMP-2, 9, 11 and 13 and TIMP-3 were evaluated by immunohistochemistry in the tumor area from previous negative biopsies and RP. RESULTS: Immunohistochemical staining values (Score) for MMPs (-11 and -13) and TIMP-3 showed no significant differences when comparing the areas of negative biopsies where tumors subsequently developed with those of the RP. However, we did observe a significant difference in the increased expression of MMP-2 (P=.002) and MMP-9 (P=.001) in the tumor area of the RP with respect to the corresponding area of the previous negative biopsy. CONCLUSIONS: Our data indicate a higher overall expression of MMP-2 and MMP-9 in the tumor area of the RP compared to the corresponding areas of the negative previous biopsy, which seems to be associated to the process of malignant transformation.

miR-9-5p attenuates ischemic stroke through targeting ERMP1-mediated endoplasmic reticulum stress.

Ischemic stroke (IS) is a cerebrovascular disease with serious neurological function impairment, which may activate endoplasmic reticulum (ER) stress. However, the underlying regulatory mechanism of ER stress under IS remains unclear. miR-9-5p is enriched in the brain tissues and plays a role in the pathological process of IS. Therefore, the purpose of this study is to explore the effect of miR-9 on ER stress and underlying mechanism in IS. Here, a middle cerebral artery occlusion (MCAO) rat model was utilized to examine the alteration of brain pathology, and the expressions of miR-9 and ER stress-related proteins. Then SH-SY5Y cells with oxygen-glucose deprivation (OGD) were performed to further evaluate the functional role of miR-9 and preliminary mechanism. The results showed that miR-9 levels were decreased in the ischemic region of rats after MCAO. MCAO significantly increased the brain infract volume, reduced Nissl bodies and cell apoptosis, and increased ER stress-related proteins (ERMP1, GRP78, p-PERK, p-eIF2α and CHOP). Furthermore, overexpression of miR-9 by miR-9 mimics increased cell viability, inhibited LDH activity and cell apoptosis, and inactivated ER stress in OGD-neurons. Luciferase activity results showed that miR-9 negatively regulated ERMP1 expression by directly targeting ERMP1 3' UTR. Subsequently, we found that ERMP1 overexpression reversed the inhibition of miR-9 on GRP78-PERK-CHOP pathway in OGD neurons. In summary, our results suggest that the attenuation of miR-9 on ischemic injury may be involved in targeting ERMP1-mediated ER stress, which provides an available target for IS treatment.

Venoms of Iranian Scorpions (Arachnida, Scorpiones) and Their Potential for Drug Discovery.

Scorpions, a characteristic group of arthropods, are among the earliest diverging arachnids, dating back almost 440 million years. One of the many interesting aspects of scorpions is that they have venom arsenals for capturing prey and defending against predators, which may play a critical role in their evolutionary success. Unfortunately, however, scorpion envenomation represents a serious health problem in several countries, including Iran. Iran is acknowledged as an area with a high richness of scorpion species and families. The diversity of the scorpion fauna in Iran is the subject of this review, in which we report a total of 78 species and subspecies in 19 genera and four families. We also list some of the toxins or genes studied from five species, including Androctonus crassicauda, Hottentotta zagrosensis, Mesobuthus phillipsi, Odontobuthus doriae, and Hemiscorpius lepturus, in the Buthidae and Hemiscorpiidae families. Lastly, we review the diverse functions of typical toxins from the Iranian scorpion species, including their medical applications.

Production, Partial Purification, and Biochemical Characterization of a Thermotolerant Alkaline Metallo-protease from Staphylococcus sciuri.

Protease-producing Staphylococcus sciuri was isolated from poultry soil samples and culture conditions for protease production were optimized. The isolated protease showed a maximum activity of 235.1 U/ml. Enzyme purification procedure involved ammonium sulphate precipitation and Sephacryl S-200 HR gel filtration chromatography (GFC). The purification process resulted in the production of three protease fractions namely protease І (metallo-alkaline protease), II, and IІІ. The metallo-alkaline protease was purified to 25.49-fold with specific activity of 982.22 U/mg and 3.76% recovery. The partially purified metallo-protease was optimally active at pH 10.0 and 70 °C and exhibited thermal stability up to 50 °C. The protease activity was enhanced by Ca2+ and Mg2+, completely inhibited by Hg2+ and Cu2+, and significantly reduced by EDTA. The protease showed significant stability towards various surfactants, including SDS. The Km and Vmax values were 0.68 mg/ml and 166.66 nmol of azocasein/ml/h, respectively, while the activation energy (Ea) was 3.07 Kcal/mol. Hence, it is evident that the produced protease possesses unique characteristics and could be a plausible candidate for various industrial and biotechnological applications.

A sigma factor RpoD negatively regulates temperature-dependent metalloprotease expression in a pathogenic Vibrio splendidus.

Vibrio splendidus is an important aquatic pathogen that can cause typical symptoms of skin ulceration syndrome (SUS) in sea cucumber Apostichopus japonicus. A metalloprotease Vsm from V. splendidus, has been reported to be an important virulence factor for SUS outbreak. In the present study, the mRNA expression level of vsm was found to correlate to temperature with a peak expression at 28 °C. In contrast, the expression of a sigma factor rpoD, was significantly repressed at 28 °C. A predicted RpoD binding site in the promoter region of vsm revealed the potential regulation of RpoD on vsm expression. Electrophoretic mobility shift assay showed that the purified recombinant RpoD could specifically bind to the promoter region of vsm. Co-transfection of vsm promotor and pT3-rpoD into E. coli significantly inhibited the ß-Galactosidase activities in a temperature-dependent manner, and the activities were 0.41-, 0.89- and 0.18-fold at 10 °C, 28 °C and 37 °C compared to the control DH5α/pT3. A rpoD overexpression strain Vs/JRTcrpoD was constructed to further examine the effect of RpoD on the expression of vsm in vivo. By real time RT-PCR analysis, vsm expression level was 0.47-fold in Vs/JRTcrpoD compared to that in Vs/JRTc. Consistently, the metalloprotease activities in Vs/JRTcrpoD was decreased by 18% compared to that in Vs/JRTc. All the results suggested that the sigma factor RpoD, showed a negative regulation on expression of vsm gene by directly interacting with the promoter region of vsm.

L1CAM induces perineural invasion of pancreas cancer cells by upregulation of metalloproteinase expression.

Pancreas cancer cells have a tendency to invade along nerves. Such cancerous nerve invasion (CNI) is associated with poor outcome; however, the exact mechanism that drives cancer cells to disseminate along nerves is unknown. Immunohistochemical analysis of human pancreatic ductal adenocarcinoma (PDAC) specimens showed overexpression of the L1 cell adhesion molecule (L1CAM) in cancer cells and in adjacent Schwann cells (SC) in invaded nerves. By modeling the neural microenvironment, we found that L1CAM secreted from SCs acts as a strong chemoattractant to cancer cells, through activation of MAP kinase signaling. L1CAM also upregulated expression of metalloproteinase-2 (MMP-2) and MMP-9 by PDAC cells, through STAT3 activation. Using a transgenic Pdx-1-Cre/KrasG12D /p53R172H (KPC) mouse model, we show that treatment with anti-L1CAM Ab significantly reduces CNI in vivo. We provide evidence of a paracrine response between SCs and cancer cells in the neural niche, which promotes cancer invasion via L1CAM secretion.

csrB Gene Duplication Drives the Evolution of Redundant Regulatory Pathways Controlling Expression of the Major Toxic Secreted Metalloproteases in Vibrio tasmaniensis LGP32.

CsrBs are bacterial highly conserved and multiple-copy noncoding small RNAs (sRNAs) that play major roles in cell physiology and virulence. In the Vibrio genus, they are known to be regulated by the two-component system VarS/VarA. They modulate the well-characterized quorum sensing pathway controlling virulence and luminescence in Vibrio cholerae and Vibrio harveyi, respectively. Remarkably, Vibrio tasmaniensis LGP32, an oyster pathogen that belongs to the Splendidus clade, was found to have four copies of csrB, named csrB1-4, compared to two to three copies in other Vibrio species. Here, we show that the extra csrB4 copy results from a csrB3 gene duplication, a characteristic of the Splendidus clade. Interestingly, csrB genes are regulated in different ways in V. tasmaniensis, with csrB1 expression being independent of the VarS/VarA system. We found that a complex regulatory network involving CsrBs, quorum sensing, and the stationary-phase sigma factor σS redundantly but differentially controls the production of two secreted metalloproteases, Vsm and PrtV, the former being a major determinant of the V. tasmaniensis extracellular product toxicity. In particular, we identified a novel VarS/VarA-dependent but CsrB-independent pathway that controls positively both Vsm production and PrtV production as well as rpoS expression. Altogether, our data show that a csrB gene duplication event in V. tasmaniensis supported the evolution of the regulatory network controlling the expression of major toxic secreted metalloproteases, thereby increasing redundancy and enabling the integration of additional input signals.IMPORTANCE The conserved CsrB sRNAs are an example of sibling sRNAs, i.e., sRNAs which are present in multiple copies in genomes. This report illustrates how new copies arise through gene duplication events and highlights two evolutionary advantages of having such multiple copies: differential regulation of the multiple copies allows integration of different input signals into the regulatory network of which they are parts, and the high redundancy that they provide confers a strong robustness to the system.

Expression levels of A disintegrin and metalloproteases (ADAMs), and Th17-related cytokines and their association with Helicobacter pylori infection in patients with gastroduodenal diseases.

Expression levels of A disintegrin and metalloproteases (ADAMs) (10 and 17) and Th17-related cytokines [interleukin (IL) 17A, IL-17F, IL-33, IL-23, IL-23R] were investigated by quantitative real time polymerase chain reaction in gastric biopsies of patients with different gastroduodenal pathologies in the presence and absence of Helicobacter pylori infection. Patients with gastric cancer (GC) (n = 70, intestinal-type 38 and diffuse type 32), peptic ulcer disease [n = 50, duodenal ulcer (DU) 16 and gastric ulcer (GU) 34] and functional dyspepsia (n = 120) were included in the study. Further, the expression levels of ADAMs and Th17 cytokines were correlated with H. pylori cytotoxin-associated genes pathogenicity island (cagPAI) status. Expression levels of ADAMs (10 and 17) and Th17-related cytokines (IL-17A, IL-23, IL-23R) were significantly higher in H. pylori-positive than in H. pylori-negative gastric biopsies. Significant increase in ADAM17 and Th17 cytokines (IL-17A and IL-23) expressions was observed in patients with GU and intestinal-type GC in the presence of H. pylori infection and in strains harbouring intact cagPAI. Expression levels of IL-17A, IL-23 and ADAM17 were strongly correlated with GU and intestinal-type GC and weakly with DU and diffuse-type GC in the presence of H. pylori infection. Higher expression levels of ADAM17 and Th17 cytokines (IL-17A and IL-23), and their strong correlation with GU and intestinal-type GC patients in the presence of H. pylori and its intact cagPAI status, suggest a possible role of strain specificity in the pathogenesis of these diseases.

Abietic acid attenuates IL-1ß-induced inflammation in human osteoarthritis chondrocytes.

Abietic acid has been reported to have anti-inflammatory activity. However, whether abietic acid has anti-inflammatory effects against osteoarthritis (OA) remains unclear. The present study aimed to measure the anti-inflammatory effects of abietic acid on OA in vitro. Human osteoarthritis chondrocytes were pretreated with abietic acid 1 h before IL-1ß treatment. The results showed that treatment of abietic acid significantly inhibited IL-1ß-induced TNF-α, NO, PGE2 production, and COX-2 expression. Abietic acid also concentration-dependently suppressed MMP1, MMP3, and MMP13 production induced by IL-1ß. Moreover, the increased phosphorylation levels of NF-κB p65 and IκBα were inhibited by the treatment of abietic acid. We also found that the expression of PPAR-γ was increased by abietic acid. The inhibition of abietic acid on TNF-α, NO, and PGE2 production were reversed by GW9662, the inhibitor of PPAR-γ. In conclusion, the study elucidated abietic acid suppressed IL-1ß-induced inflammation in human osteoarthritis chondrocytes by activating PPAR-γ.

The extracellular proteases produced by Vibrio parahaemolyticus.

Vibrio parahaemolyticus, a Gram-negative bacterium, inhabits marine and estuarine environments and it is a major pathogen responsible globally for most cases of seafood-associated gastroenteritis in humans and acute hepatopancreatic necrosis syndrome in shrimps. There has been a dramatic worldwide increase in V. parahaemolyticus infections over the last two decades. The pathogenicity of V. parahaemolyticus has been linked to the expression of different kinds of virulence factors including extracellular proteases, such as metalloproteases and serine proteases. V. parahaemolyticus expresses the metalloproteases; PrtV, VppC, VPM and the serine proteases; VPP1/Protease A, VpSP37, PrtA. Extracellular proteases have been identified as potential virulence factors which directly digest many kinds of host proteins or indirectly are involved in the processing of other toxic protein factors. This review summarizes findings on the metalloproteases and serine proteases produced by V. parahaemolyticus and their roles in infections. Identifying the role of V. parahaemolyticus virulence-associated extracellular proteases deepens our understanding of diseases caused by this bacterium.

Bothrops jararaca accessory venom gland is an ancillary source of toxins to the snake.

In Viperidae snakes, it has been attributed to the main venom gland, a component of the venom gland apparatus, the function of synthesizing all venom toxins and storing them inside a basal-central lumen. However, the role of the accessory gland is still unknown. Here, we analyzed the proteome and the transcriptome of the accessory gland during venom production and secretion cycle. We showed that the accessory gland expresses and synthesizes toxins that are similar to those produced by the main venom gland such as C-type lectin/C-type lectin-like proteins, metalloproteinase, phospholipase A2, cysteine rich secretory protein, nerve growth factor, vascular endothelial growth factor, serine proteinase, and l-amino acid oxidase. Our data have shown that toxin synthesis in the accessory gland is asynchronous when compared to the same process in the venom gland. Moreover, this gland also expresses inhibitors of venom phospholipases A2 and metalloproteinases. Transcriptome analysis showed that the transcripts that correspond to toxins in the accessory gland have a good correlation to the main venom gland transcripts. Therefore, it is proposed that the accessory gland is an ancillary source of toxins to the snake, and provides inhibitors that could control venom toxicity (and integrity) during storage. SIGNIFICANCE: In this study, we propose that the accessory venom gland acts as an important ancillary source of toxins to the snake, in lieu of a depleted main venom gland, and provides inhibiting agents that control venom toxicity (and integrity) during its storage.

Proteomic Analyses of the Unexplored Sea Anemone Bunodactis verrucosa.

Cnidarian toxic products, particularly peptide toxins, constitute a promising target for biomedicine research. Indeed, cnidarians are considered as the largest phylum of generally toxic animals. However, research on peptides and toxins of sea anemones is still limited. Moreover, most of the toxins from sea anemones have been discovered by classical purification approaches. Recently, high-throughput methodologies have been used for this purpose but in other Phyla. Hence, the present work was focused on the proteomic analyses of whole-body extract from the unexplored sea anemone Bunodactis verrucosa. The proteomic analyses applied were based on two methods: two-dimensional gel electrophoresis combined with MALDI-TOF/TOF and shotgun proteomic approach. In total, 413 proteins were identified, but only eight proteins were identified from gel-based analyses. Such proteins are mainly involved in basal metabolism and biosynthesis of antibiotics as the most relevant pathways. In addition, some putative toxins including metalloproteinases and neurotoxins were also identified. These findings reinforce the significance of the production of antimicrobial compounds and toxins by sea anemones, which play a significant role in defense and feeding. In general, the present study provides the first proteome map of the sea anemone B. verrucosa stablishing a reference for future studies in the discovery of new compounds.

Contribution of photodynamic therapy in wound healing: A systematic review.

OBJECTIVE: We researched articles that used photodynamic therapy (PDT) for skin wound healing in humans. METHODS: The systematic review was conducted through scientific articles that investigated the action of PDT on wound healing in humans, published from July 2005 to March 2017, in the data bases PubMed and LILACS. RESULTS: The main types of wound described in selected articles in this review were chronic ulcer and non-melanoma skin cancer. For accomplishing the PDT, second generation of photosensitizing agents with laser or light emitting diode were used. The studies demonstrated that PDT contribute in several ways to the wound healing process: leading to cellular death; reducing or increasing inflammation; stimulating fibroblasts proliferation and, consequently, of collagen and elastin; raising transforming growth factor beta and metalloproteinases. Based on this, PDT provided good results in wound healing process, acting in several steps and accelerating tissue repair. CONCLUSIONS: PDT improved healing in many wound models in humans, revealing itself as a promising therapeutic modality for stimulating wound healing and remodelling.

Estrogen receptor α protects pancreatic ß-cells from apoptosis by preserving mitochondrial function and suppressing endoplasmic reticulum stress.

Estrogen receptor α (ERα) action plays an important role in pancreatic ß-cell function and survival; thus, it is considered a potential therapeutic target for the treatment of type 2 diabetes in women. However, the mechanisms underlying the protective effects of ERα remain unclear. Because ERα regulates mitochondrial metabolism in other cell types, we hypothesized that ERα may act to preserve insulin secretion and promote ß-cell survival by regulating mitochondrial-endoplasmic reticulum (EndoRetic) function. We tested this hypothesis using pancreatic islet-specific ERα knockout (PERαKO) mice and Min6 ß-cells in culture with Esr1 knockdown (KD). We found that Esr1-KD promoted reactive oxygen species production that associated with reduced fission/fusion dynamics and impaired mitophagy. Electron microscopy showed mitochondrial enlargement and a pro-fusion phenotype. Mitochondrial cristae and endoplasmic reticulum were dilated in Esr1-KD compared with ERα replete Min6 ß-cells. Increased expression of Oma1 and Chop was paralleled by increased oxygen consumption and apoptosis susceptibility in ERα-KD cells. In contrast, ERα overexpression and ligand activation reduced both Chop and Oma1 expression, likely by ERα binding to consensus estrogen-response element sites in the Oma1 and Chop promoters. Together, our findings suggest that ERα promotes ß-cell survival and insulin secretion through maintenance of mitochondrial fission/fusion-mitophagy dynamics and EndoRetic function, in part by Oma1 and Chop repression.

Expression, purification and characterization of two leucine aminopeptidases of the blood fluke, Schistosoma mansoni.

Schistosomiasis is a major neglected tropical disease (NTD) and considered the most important of the human helminthiases in terms of morbidity and mortality. Whereas treatment with praziquantel has been effective since the 1980s, the potential for the emergence of drug resistance has propelled the search for new interventions. Studies have revealed key roles of proteases in parasitic helminths during establishment of infection, tissue invasion, immune evasion, parasite feeding and development throughout the different developmental stages, pinpointing them as possible candidates. The leucine aminopeptidases (LAPs), members of the M17 family of Zn-metalloproteases, preferentially cleave leucine (Leu) residues at the N-terminal end of proteins and short peptides. These enzymes display broad proteolytic activities beyond Leu hydrolysis and are involved in processing, maturation, activation and/or degradation of substrates. As a vaccine immunogen, LAP induces protection against infection with the liver fluke Fasciola hepatica. Herein, two LAPs, SmLAP1 (Smp_030000) and SmLAP2 (Smp_083870) of the human blood fluke Schistosoma mansoni were cloned, expressed, purified and biochemically characterized. The enzymes differed in activity against diagnostic substrates, including leucine, methionine and arginine, with an optimal pH of 8.0. The activity increased in the presence of Mg+2 and Mn+2, and was inhibited by bestatin, a specific inhibitor of aminopeptidase. In addition, 1,10-phenanthroline and EDTA inhibited the enzymatic activity of SmLAP2. Finally, immunolocalization using antibodies specific for SmLAP1 and SmLAP2 identified the expression of these proteases in the egg and adult developmental stages of S. mansoni, and in intestinal epithelia, vitelline cells and sub-tegumental regions of the parasite. Characterization of schistosome proteases not only enhances understanding of the biology of schistosomes and schistosomiasis, but may also provide novel intervention approaches.

Human pericytes adopt myofibroblast properties in the microenvironment of the IPF lung.

Idiopathic pulmonary fibrosis (IPF) is a fatal disease of unknown etiology characterized by a compositionally and mechanically altered extracellular matrix. Poor understanding of the origin of α-smooth muscle actin (α-SMA) expressing myofibroblasts has hindered curative therapies. Though proposed as a source of myofibroblasts in mammalian tissues, identification of microvascular pericytes (PC) as contributors to α-SMA-expressing populations in human IPF and the mechanisms driving this accumulation remain unexplored. Here, we demonstrate enhanced detection of α-SMA+ cells coexpressing the PC marker neural/glial antigen 2 in the human IPF lung. Isolated human PC cultured on decellularized IPF lung matrices adopt expression of α-SMA, demonstrating that these cells undergo phenotypic transition in response to direct contact with the extracellular matrix (ECM) of the fibrotic human lung. Using potentially novel human lung-conjugated hydrogels with tunable mechanical properties, we decoupled PC responses to matrix composition and stiffness to show that α-SMA+ PC accumulate in a mechanosensitive manner independent of matrix composition. PC activated with TGF-ß1 remodel the normal lung matrix, increasing tissue stiffness to facilitate the emergence of α-SMA+ PC via MKL-1/MTRFA mechanotranduction. Nintedanib, a tyrosine-kinase inhibitor approved for IPF treatment, restores the elastic modulus of fibrotic lung matrices to reverse the α-SMA+ phenotype. This work furthers our understanding of the role that microvascular PC play in the evolution of IPF, describes the creation of an ex vivo platform that advances the study of fibrosis, and presents a potentially novel mode of action for a commonly used antifibrotic therapy that has great relevance for human disease.

Regulation systems of protease and hemolysin production in Vibrio vulnificus.

Vibrio vulnificus, a gram-negative halophilic estuarine bacterium, is an opportunistic human pathogen that causes rapidly progressive fatal septicemia and necrotizing wound infection. This species also causes hemorrhagic septicemia called vibriosis in cultured eels. It has been proposed that a range of virulence factors play roles in pathogenesis during human and/or eel infection. Among these factors, a metalloprotease (V. vulnificus protease [VVP]) and a cytolytic toxin (V. vulnificus hemolysin [VVH]) are of significant importance. VVP elicits the characteristic edematous and hemorrhagic skin damage, whereas VVH exhibits powerful hemolytic and cytolytic activities and contributes to bacterial invasion from the intestine to the blood stream. In addition, a few V. vulnificus strains isolated from diseased eels have recently been found to produce a serine protease designated as V. vulnificus serine protease (VvsA) instead of VVP. Similarly to VVP, VvsA may possess various toxic activities such as collagenolytic, cytotoxic and edema-forming activity. In this review, regulation of V. vulnificus VVP, VVH and VvsA is clarified in terms of expression at the mRNA and protein levels. The explanation is given on the basis of the quorum sensing system, which is dependent on bacterial cell density. In addition, the roles of environmental factors and global regulators, such as histone-like nucleoid structuring protein, cyclic adeno monophosphate receptor protein, RpoS, HlyU, Fur, ToxRS, AphB and LeuO, in this regulation are outlined. The cumulative impact of these regulatory systems on the pathogenicity of V. vulnificus is here delineated.