Reduction of myocardial infarct size by doxycycline: A role for plasmin inhibition

Myocardial ischemia-reperfusion (I/R) is associated with the activation of matrix metalloproteinases (MMPs) and serine proteases. We hypothesized that activation of MMPs and the serine protease plasmin contribute to early cardiac myocyte death following I/R and that broad-spectrum protease inhibition with doxycycline (DOX) preserves myocyte viability. Rats treated daily with or without DOX beginning 48 h prior to experimentation were subjected to 30 min of coronary occlusion and 2 days of reperfusion. DOX pre-treatment reduced infarct size by 37%. DOX attenuated increases in MMP-9 and plasmin levels as determined by gelatin zymography and immunoblot, respectively. Neutrophil extravasation was unaltered by DOX as assessed by myeloperoxidase (MPO) activity. To examine the contribution of MMP-9 and plasmin to myocyte injury, cultures of neonatal rat ventricular myocytes (NRVMs) were treated for 48 h with 83 kDa MMP-9 or plasminogen in the presence or absence of DOX. MMP-9 treatment did not affect myocyte viability. Plasminogen treatment led to increased plasmin activity, resulting in loss of ₁-integrin, NRVM detachment and apoptosis. DOX co-treatment inhibited plasmin activity and preserved NRVM attachment, whereas co-treatment with the broad-spectrum MMP inhibitor GM6001 had no effect. These results indicate that plasmin causes disruption of myocyte attachment and viability independently of MMP activation in vitro and that inhibition of plasmin by DOX may reduce I/R-induced myocyte death in vivo through the inhibition of plasmin. (Mol Cell Biochem 270: 1–11, 2005)     

The effect of VDR gene polymorphisms and vitamin D level on blood pressure,risk of preeclampsia,gestational age,and body mass index

We investigated the influence of vitamin D receptor (VDR) polymorphisms and vitamin D level on the blood pressure and the risk of preeclampsia. In a case-control study, 200 pregnant women, including 100 individuals with preeclampsia along with 100 healthy pregnant women, were studied for VDR FokI, TaqI, and BmsI polymorphisms and serum 25 (OH)-D level using polymerase chain reaction-restriction fragment length polymorphism method and commercial kit, respectively. The mean level of 25 (OH)-D in preeclamptic patients was significantly lower (16.6 ± 4.2 ng/mL, P < 0.001) compared with controls (19.6 ± 3.8 ng/mL). Among all women, a significantly higher systolic blood pressure and before-pregnancy body mass index and also lower gestational age were observed in the presence of 25 (OH)-D level < 20 ng/mL compared with the 20 to 30 ng/mL. A significantly higher frequency of VDR FokI C allele in preeclamptic patients (83%) than controls (74%) was associated with a 1.72-fold increased risk of preeclampsia. In all the studied individuals, the systolic and diastolic blood pressures were significantly higher in the presence of the FokI CC genotype compared with the TC and TT+TC genotypes. Neither VDR Taq1 nor VDR BmsI was associated with the risk of preeclampsia. The haplotype FokI C, TaqI C and BmsI A (CCA) compared with haplotype CTG increased the risk of preeclampsia by 1.4-fold (P = 0.33). Our study suggests an association between VDR FokI polymorphism and an insufficient serum level of 25 (OH)-D with the risk of preeclampsia and also the influence of insufficient 25 (OH)-D level and VDR FokI polymorphism on maternal factors, including blood pressure.     

Ionenkanalerkrankungen des Gehirns – monogene Epilepsien

Epilepsy is one of the most common neurological disorders and is characterized by recurrent, unprovoked epileptic seizures. Seizures are generated by spontaneous, synchronous neuronal discharges which induce disturbances of perception or behaviour. About one third of all epilepsies are primarily caused by genetic factors. These so-called idiopathic epilepsies occur without observable structural alterations in the brain. Mutations in genes encoding neuronal ion channels play a central role in the etiology of such epilepsies. In this review, mutations in ion channel genes associated with idiopathic epilepsies and their functional consequences are described. The underlying pathomechanisms and consequences for treatment are discussed.     

Effects of the utilization of homeopathic elements in commercial diluent on swine sperm viability

It has been speculated that the homeopathic treatment of sperm cells in order to improve semen quality could be promising. However, few data is available and its use in spermatozoa requires investigation. It is well established that mitochondrial membrane potential is an important viability parameter of spermatozoa and it is intimately related to reproductive efficiency. In this manner, new technologies in order to improve the activity of sperm cells and, finally, the fecundity of swine herds are of extremely importance. Due to the lack of knowledge of homeopathic treatment effect on spermatozoa, the aim of the present study was to verify the effect of three different homeopathic treatments on viability of boar sperm cells. Three homeopathic treatments composed by Pulsatila CH6, Pulsatila and Avena CH6, Avena CH6 and one control treatment (sucrose) were added to diluted boar semen, which were cooled for 24 or 48 h. Interestingly, no positive effect of homeopathic treatments was observed over semen viability. However, it was demonstrated that the 24 h of cooling storage provided more viable sperm cells when compared to the 48-h period. This effect of storage period on sperm viability was assessed by intact plasmatic membrane, intact acrosome and mitochondrial membrane potential evaluation.     

Activin A is essential for Feeder‐free culture of human induced pluripotent stem cells

Feeder‐free culture of human induced pluripotent stem (hiPS) cells is necessary for their clinical application to avoid adverse effects of foreign proteins. hiPS cells were cultured with combinations of activin (A), CHIR99021 (C), basic fibroblast growth factor (F), and leukemia inhibitory factor (L) under feeder‐free conditions. Culture was terminated after 12 passages or when the cell morphology changed from pluripotency. Pluripotency was analyzed by alkaline phosphatase (ALP) staining and immunostaining with antibodies to Oct3/4, Nanog, SSEA4, and TRA‐1‐60. SB431542 (SB), an activin inhibitor, was added to the culture, and the morphology of the cells was observed. hiPS cells cultured with A, AC, and ACL after 12 passages were positive for ALP staining. Oct3/4 was positive in hiPS cells cultured with A, AC, and ACL. hiPS cells were positive for Nanog when cultured with A and AC; however, Nanog signal was weaker in cells cultured with ACL. SSEA4 was positive in hiPS cells cultured with A and AC but almost negative in those cultured with ACL. hiPS cells were positive for TRA‐1‐60 when cultured with A, AC, and ACL. hiPS cells lose their undifferentiated morphology at six passages when cultured with A + SB, five passages with AC + SB, and nine passages with ACL. We conclude that feeder‐free culture of hiPS cells requires A or AC to maintain pluripotency. J. Cell. Biochem. 114: 584–588, 2013. © 2012 Wiley Periodicals, Inc.     

A novel tumor metastasis suppressor gene LASS2/TMSG1 interacts with vacuolar ATPase through its homeodomain

LASS2/TMSG1 was a novel tumor metastasis suppressor gene, which was first cloned by our laboratory from non‐metastatic and metastatic cancer cell variants of human prostate carcinoma PC‐3M using mRNA differential display in 1999. LASS2/TMSG1 could interact with the C subunit of vacuolar ATPase (V‐ATPase, ATP6V0C) and regulate V‐ATPase activity. In an attempt to provide molecular mechanism of the interaction between LASS2/TMSG1 and V‐ATPase, we constructed four variant transfectants containing different functional domain of LASS2/TMSG1 and stably transfected the variants to human prostate cancer cell line PC‐3M‐1E8 cell with high metastatic potential. Results showed that there were no obvious differences of V‐ATPase expression among different transfected cells and the control. However, V‐ATPase activity and intracellular pH was significantly higher in the variant transfectants with Homeodomain of LASS2/TMSG1 than that in the control using the pH‐dependent fluorescence probe BECEF/AM. Immunoprecipitation, immunofluorescence and immuno‐electron microscope alone or in combination demonstrated the direct interaction of Homeodomain of LASS2/TMSG1 and ATP6V0C. Loss of Homeodomain markedly enhanced the proliferation ability but weakened the apoptotic effect of LASS2/TMSG1 in PC‐3M‐1E8 cells. These lines of results for the first time contribute to the conclusion that LASS2/TMSG1 could regulate V‐ATPase activity and intracellular pH through the direct interaction of its Homeodomain and the C subunit of V‐ATPase. Their interaction could play important roles in the apoptosis of tumor cells. J. Cell. Biochem. 114: 570–583, 2013. © 2012 Wiley Periodicals, Inc.