Effect of Kang Fu Yan capsule on phenol mucilage-induced intrauterine adhesion injury in female rats

Purpose: To investigate the effect of Kang fu yan capsule (KFYC) on phenol mucilage-induced intrauterine adhesion (IUA) in a rat model, and the underlying mechanisms. Methods: An IUA model was established by injecting 0.06 mL of 25 % phenol mucilage into the uterus of female Sprague-Dawley rats. The IUA model rats (n=59) were randomly divided into 5 groups: IUA group, fuke qianjin tablet group (FKQJT, 0.22 mg/kg), and 3 KFYC groups given different doses of the drug i.e. 0.13, 0.39and 1.17 mg/kg. A group of 10 healthy female rats served as control. After 19 days treatment, blood samples were collected for determination of IL-2 and IL-10 by ELISA, while uterine tissues were subjected to histological examination using hematoxylin and eosin staining (H&E) and Masson staining. Expressions of Notch1, recombination signal binding protein-JK (RBP-JK), a disintegrin and metalloprotease (ADAM)-12, ADAM-15, matrix metalloprotein-9 (MMP-9), and inhibitor of NF-κB (IĸB) in uterine tissues were determined using RT-qPCR and western blot analysis. Results: Compared to IUA group, histological results showed reduced inflammatory cell infiltration in rat uterine tissue of KFYC group. Moreover, KFYC significantly reversed uterine fibrosis (p < 0.05). Serum concentrations of IL-2 significantly decreased in KFYC groups (p < 0.05 or p < 0.01), and there was significant increases the serum concentrations of IL-10 in KFYC groups (p < 0.05 or < 0.01), when compared to IUA group. The mRNA and protein expressions of Notch1, RBP-JK, ADAM-12, ADAM-15, MMP-9 were also significantly down-regulated (p < 0.05), while protein expression of IĸB was upregulated in KFYC group, when compared to IUA group. Conclusion: KFYC exerts an anti-IUA effect via amelioration of uterine inflammation and fibrosis, probably via a mechanism involving regulation of Notch1/ADAM pathway.

Simvastatin impairs myoblast proliferation and myotube formation of C2C12 myoblasts and in mouse skeletal muscle

Statins reduce cardiovascular complications in patients with high LDL-cholesterol but are associated with myopathy. We investigated the possibility that statins impair skeletal muscle regeneration by assessing simvastatin toxicity on C2C12 myoblasts and myotubes and mouse skeletal muscle. Simvastatin increased plasma membrane permeability and decreased the cellular ATP content in both myoblasts and myotubes, but with a stronger effect on myoblasts. While insulin prevented cytotoxicity up to 8 hours after addition of simvastatin to myotubes, prevention in myoblasts required simultaneous addition. Mevalonate and geranylgeraniol also prevented simvastatin-associated cytotoxicity on myoblasts and myotubes. Simvastatin impaired the phosphorylation of the insulin receptor (IR β), Akt ser473 and S6rp, and increased phosphorylation of AMPK thr172 in both myotubes and myoblasts, which was prevented by insulin and mevalonate. Simvastatin impaired oxygen consumption and increased superoxide production by myoblasts and myotubes and induced apoptosis via cytochromc c release. In addition, simvastatin impaired proliferation and fusion of myoblasts to myotubes by inhibiting the expression of the nuclear transcription factor MyoD and of the metalloprotease ADAM-12. Decreased expression of the proliferation factor Ki-67 and of ADAM-12 were also observed in gastrocnemius of mice treated with simvastatin. In conclusion, myoblasts were more susceptible to the toxic effects of simvastatin and simvastatin impaired myoblast proliferation and myotube formation. Impaired muscle regeneration represents a new mechanism of statin myotoxicity and may be important in statin-associated myopathy.

A disintegrin and metalloproteinase 12 prevents heart failure by regulating cardiac hypertrophy and fibrosis

A disintegrin and metalloproteinase (ADAM)12 is considered to promote cardiac dysfunction based on the finding that a small-molecule ADAM12 inhibitor, KB-R7785, ameliorated cardiac function in a transverse aortic constriction (TAC) model by inhibiting the proteolytic activation of heparin-binding-EGF signaling. However, this compound has poor selectivity for ADAM12, and the role of ADAM12 in cardiac dysfunction has not yet been investigated using genetic loss-of-function mice. We revealed that ADAM12 knockout mice showed significantly more advanced cardiac hypertrophy and higher mortality rates than wild-type mice 4 wk after TAC surgery. An ADAM12 deficiency resulted in significantly more expanded cardiac fibrosis accompanied by increased collagen-related gene expression in failing hearts. The results of a genome-wide transcriptional analysis suggested a strongly enhanced focal adhesion- and fibrosis-related signaling pathway in ADAM12 knockout hearts. The loss of ADAM12 increased the abundance of the integrinβ1 subunit and transforming growth factor (TGF)-β receptor types I and III, and this was followed by the phosphorylation of focal adhesion kinase, Akt, mammalian target of rapamycin, ERK, and Smad2/3 in the heart, which resulted in cardiac dysfunction. The present results revealed that the loss of ADAM12 enhanced focal adhesion and canonical TGF-β signaling by regulating the abundance of the integrinβ1 and TGF-β receptors. NEW & NOTEWORTHY In contrast to a long-believed cardio-damaging role of a disintegrin and metalloproteinase (ADAM)12, cardiac hypertrophy was more severe, cardiac function was lower, and mortality was higher in ADAM12 knockout mice than in wild-type mice after transverse aortic constriction surgery. The loss of ADAM12 enhanced focal adhesion- and fibrosis-related signaling pathways in the heart, which may compromise cardiac function. These results provide insights for the development of novel therapeutics that target ADAM12 to treat heart failure.

Predição de crescimento intrauterino restrito no rastreio combinado de primeiro trimestre

Introdução: O conceito de restrição do crescimento fetal tardio (CFT) foi introduzido para descrever fetos com insuficiência placentária que não é grave o suficiente para resultar em CFT com fluxo arterial umbilical anormal, mas é grave o suficiente para associar-se com alto risco perinatal ou complicações de longo prazo. Entretanto, há necessidade de retroceder a investigação para o primeiro trimestre, na esperança de possível intervenção. Objetivo: Realizar revisão bibliográfica do tema. Material e Métodos: As palavras-chave CIUR (crescimento intrauterino restrito) e IUGR (intrauterine growth restriction) foram identificadas na Biblioteca Virtual em Saúde (BVS). O termo CIUR também foi pesquisado na Literatura Latino-americana e do Caribe em Ciências da Saúde (LILACS), nos últimos dez anos. Com o auxílio do Endnote, foram realizadas pesquisas no PubMed usando a palavra-chave (MeSH term) intrauterine growth restriction (IUGR), no período 2011‒2020. Os artigos foram baixados e foi montada uma pasta com eles em formato pdf. Posteriormente, foram importados pelo Endnote. Resultados: Na LILACS, nos últimos 10 anos, foram recuperados 20 artigos com texto completo. No PubMed, no período 2011‒2020 (até agosto), foram identificadas 5.551 referências sob o título “intrauterine growth restriction”. Utilizando como filtro a expressão “first trimester”, restaram 370 referências, e após um segundo filtro (“screening”), 308 referências. Destas, conseguimos os textos completos em .pdf de 101, que foram utilizados para a presente revisão. Modelos foram identificados incluindo dados clínicos maternos (método de contracepção, tabagismo, altura materna), métodos de imagem (Doppler da artéria uterina, avaliando o índice de pulsatilidade — PI) e avaliação bioquímica (fator de crescimento placentário — PlGF, gonadotrofina coriônica — βhCG, pregnancy-associated plasma protein-A — PAPP-A, soluble fms-like tytosine kinase — sFlt-1 e disintegrin and metalloprotease — ADAM 12). Conclusão: Neste estudo, foram utilizados diferentes modelos com associação dos fatores mencionados anteriormente. Ainda não se identificou o modelo ideal, necessitando de novos estudos.

HIF-1α is Overexpressed in Odontogenic Keratocyst Suggesting Activation of HIF-1α and NOTCH1 Signaling Pathways

Background: The odontogenic keratocyst (OKC) is an odontogenic cyst that shows aggressive and intriguing biological behavior. It is suggested that a hypoxic environment occurs in OKC, which led us to investigate the immunoexpression and location of hypoxia-inducible factor 1-alpha (HIF-1α) and other hypoxia-related proteins. Methods: Twenty cases of OKC were evaluated for the expression of Notch homolog 1 (NOTCH1), HIF-1α, disintegrin and metalloproteinase domain-containing protein 12 (ADAM-12), and heparin-binding epidermal growth factor-like growth factor (HBEGF) by immunohistochemistry and compared to eight control cases of calcifying odontogenic cystic (COC), orthokeratinized odontogenic cyst (OOC), and normal oral mucosa (OM) in basal and parabasal layers. Results: In OKC, all the proteins tested were expressed significantly higher in both basal (except for NOTCH1 and HBEGF in OOC) and suprabasal epithelial layers compared to controls. Looking at the epithelial layers within OKC, we observed an increased NOTCH1 and HIF-1α expression in parabasal layers. Conclusions: These results suggest that hypoxia occurs more intensively in OKC compared to COC, OM, and OOC. Hypoxia appeared to be stronger in parabasal layers as observed by higher HIF-1α expression in upper cells. Overexpression of NOTCH1, ADAM-12, and HBEGF in OKC was observed, which suggests that microenvironmental hypoxia could potentially regulate the expression of hypoxia-related proteins, and consequently, its clinical and biological behavior.

Certainties and uncertainties concerning the contribution of pericytes to the pathogenesis of systemic sclerosis

The role of pericytes in systemic sclerosis (SSc) is unclear because of the difficulty in phenotyping them. They are mainly distributed in the pre-capillary, capillary and post-capillary abluminal side of non-muscular micro-vessels, express platelet-derived growth factor receptors (PDGFRs), and preside over vascular integrity and regeneration. By establishing close contact with many endothelial cells, a single pericyte can regulate ion influx, mechanical stress, leukocyte diapedesis, and platelet activation. Moreover, under pathological conditions such as SSc, pericytes may acquire a contractile phenotype and respond to various stimuli, including endothelin, angiotensin II and reactive oxygen species. The pericytes of SSc patients share some molecular patterns with myofibroblasts or fibroblasts, including A disintegrin and metalloproteinase domain 12 (ADAM-12), α-smooth muscle actin (α-SMA), the extra domain A (ED-A) variant of fibronectin, and Thy-1. Following stimulation with PDGF-β or transforming growth factor-β (TGF-β), pericytes may acquire a myofibroblast phenotype, and produce extracellular matrix or indirectly promote fibroblast activation. They may also contribute to fibrosis by means of epigenetic regulation. The pericyte plasmalemma is particularly rich in caveolae containing caveolin-1, a deficit of which has been associated with defective vessel tone control and lung fibrosis in mice. Consequently, dysfunctional pericytes may underlie the microangiopathy and fibrosis observed in SSc patients. However, given its variability in biological behaviour and the lack of a pan-pericyte marker, the exact role of these cells in SSc warrants further investigation.

Abstract 500: Vitamin D Supplementation Attenuates ADAM-12-mediated Proliferation of Carotid Artery Smooth Muscle Cells of Hypercholesterolemic Swine

Introduction: Risk of hypertension, peripheral artery disease, myocardial infarction and development of human atherosclerosis has been linked to vitamin D deficiency. Atheromatous cytokines, including IL-6, TNF-α and epidermal growth factor receptor (EGFR) family growth factors are released at the site of atherosclerosis and boost the activity of proteolytic enzymes such as ADAMs (a disintegrin and metalloproteinases). ADAM-12 cleaves proHB-EGF (Heparin-binding EGF-like growth factor) activating EGFR, resulting in increased proliferation of smooth muscle cells (SMCs). The aim of this study was to examine the effect of vitamin D on IL-6 and TNF-α-induced ADAM-12, pEGFR expression, HB-EGF release and SMC proliferation. Methods: Micro-swine were fed with vitamin D-deficient high cholesterol diet, high cholesterol diet containing 900 IU of vitamin D, and high cholesterol diet containing 3000 IU of vitamin D for total of 12 months. After six months, serum cholesterol levels of 500-600 mg/dL were achieved in all the three groups. The protein expression of ADAM-12 & pEGFR, and HB-EGF release, in presence or absence of IL-6, TNF-α and Calcitriol, in SMCs was quantified by western Blot. HB-EGF release was measured by ELISA. The proliferation was assayed by [3H]-Thymidine incorporation and cell counting method. Results: The protein expression of ADAM-12 & pEGFR, HB-EGF release were significantly reduced in carotid artery SMCs isolated from Vitamin D-supplemented swine. IL-6 and TNF-α treatment increased the protein expression of ADAM-12 & pEGFR and HB-EGF release in carotid artery SMCs. Proliferation capacity was higher in SMCs isolated from Vitamin D-deficient swine carotid artery, potentiated by IL-6 and TNF-α. Calcitriol inhibited the ADAM-12, pEGFR expression and HB-EGF released in SMCs of hypercholesterolemic swine. Calcitriol also inhibited the proliferation of carotid artery SMCs isolated from Vit D-deficient, D-sufficient and D-supplemented swine. Conclusion: Together, these results suggest that vitamin D deficiency enhances proliferation of SMCs, which is potentiated by atheromatous cytokines. Whereas, vitamin D supplementation regulates ADAM-12-mediated cleavage of proHB-EGF and activation of EGFR inhibiting SMC proliferation.

Συσχέτιση του βιοχημικού παράγοντα ADAM 12 στο πρώτο τρίμηνο της κύησης με υπέρταση και καθυστέρηση της ενδομήτριας ανάπτυξης

Σκοπός: Να καθορισθεί αν ο βιοχημικός δείκτης ADAM 12 στο μητρικό αίμα, στο Α τρίμηνο της κυήσεως , δύναται να χρησιμοποιηθεί ως δείκτης για υπέρταση – προεκλαμψία και ενδομήτρια υπολλειπόμενη ανάπτυξη εμβρύου. Υλικά και μέθοδοι: Είναι συγκριτική μελέτη για τον βιοχημικό δείκτη ADAM 12 μεταξύ δύο ομάδων εγκύων .Η μία ομάδα αποτελείται από 98 έγκυες [Ν= 98 ] οι οποίες στη πορεία της κύησης ανέπτυξαν υπέρταση , προεκλαμψία και είχαν έμβρυα με ενδομήτρια υπολλειπόμενη ανάπτυξη [ IUGR , SGA- Study group]H ομάδα μαρτύρων αποτελείται από έγκυες που δεν παρουσίασαν καμία επιπλοκή στην κύηση και είχαν έμβρυα με φυσιολογική ανάπτυξη [ Control group ] .Οι δύο ομάδες είχαν όμοια περίπου δημογραφικά στοιχεία και όμοια χαρακτηριστικά Ο βιοχημικός δείκτης ADAM 12 μετρήθηκε στο μητρικό αίμα των δύο ομάδων εγκύων στο Α τρίμηνο , στις έγκυες που προσήλθαν στο τμήμα εμβρυομητρικής της Γ Μαιευτικής – Γυναικολογικής Κλινικής του Αττικού Νοσοκομείου , για προσδιορισμό κινδύνου για χρωμοσωμιακές ανωμαλίες στις 11+0 εως 13 + 6 εβδ κυήσεως .Αποτελέσματα : Δεν υπήρχε καμία στατιστική διαφορά μεταξύ του μέσου log MoMτου βιοχημικού ADAM 12 μεταξύ της ομάδας εγκύων [study group ] που ανέπτυξαν υπέρταση ,προεκλαμψία και είχαν έμβρυα με υπολλειπόμενη ανάπτυξη [SGA, IUGR]και της ομάδας εγκύων που δεν είχαν καμία επιπλοκή στη κύηση και γέννησαν φυσιοογικού βάρους νεογνά [control group ] . Αντίθετα τα επίπεδα του ADAM 12 στην υποομάδα των εγκύων που ανέπτυξαν προεκλαμψία [ PE group ] ήταν στατιστικώς χαμηλότερα σε σχέση με την ομάδα εγκύων που δεν είχε καμία επιπλοκή στην κύηση [ Control group ] .[ mean log MoM :- Ο,109vs 0,008 P=0,010].Ομοίως τα επίπεδα του βιοχημικού δείκτη ADAM 12 στο μητρικό αίμα ήταν στατιστικώς χαμηλά σε έγκυες που ανέπτυξαν προεκλαμψία και υπέρταση [PE and/orPIH] σε σχέση με τα επίπεδα ADAM 12 των εγκύων που δεν είχαν καμία επιπλοκή στην κύηση [Control group ][ mean log MoM : -0,066 vs 0,008 P=0,015].Δεν υπήρξε καμία στατιστική διαφορά στις τιμές ADAM 12 μεταξύ των εγκύων που είχαν έμβρυα με υπολλειπόμενη ανάπτυξη [ υποoμάδα SGA , IUGR group ] και της ομάδας εγκύων που δεν είχαν καμία επιπλοκή στην κύηση [ Control group ] .Συμπέρασμα : Τα επίπεδα του βιοχημικού δείκτη ADAM 12 στο α τρίμηνο της κύησης ήταν στατιστικώς χαμηλότερα σε γυναίκες που αργότερα εμφάνισαν υπέρταση και προεκλαψία στην πορεία της κύησης συγκρινόμενο με γυναίκες που δεν είχαν καμία επιπλοκή στην κύηση. Αρα ο βιοχημικός δείκτης ADAM 12 είναι χρήσιμος προγνωστικός δείκτης υπέρτασης και προεκλαμψίας .