In Vitro and In Vivo Investigation of the Angiogenic Effects of Liraglutide during Islet Transplantation

Introduction

This study investigated the angiogenic properties of liraglutide in vitro and in vivo and the mechanisms involved, with a focus on Hypoxia Inducible Factor-1α (HIF-1α) and mammalian target of rapamycin (mTOR).

Materials and Methods

Rat pancreatic islets were incubated in vitro with 10 μmol/L of liraglutide (Lira) for 12, 24 and 48 h. Islet viability was studied by fluorescein diacetate/propidium iodide staining and their function was assessed by glucose stimulation. The angiogenic effect of liraglutide was determined in vitro by the measure of vascular endothelial growth factor (VEGF) secretion using enzyme-linked immunosorbent assay and by the evaluation of VEGF and platelet-derived growth factor-α (PDGFα) expression with quantitative polymerase chain reaction technic. Then, in vitro and in vivo, angiogenic property of Lira was evaluated using immunofluorescence staining targeting the cluster of differentiation 31 (CD31). To understand angiogenic mechanisms involved by Lira, HIF-1α and mTOR activation were studied using western blotting. In vivo, islets (1000/kg body-weight) were transplanted into diabetic (streptozotocin) Lewis rats. Metabolic control was assessed for 1 month by measuring body-weight gain and fasting blood glucose.

Results

Islet viability and function were respectively preserved and enhanced (p<0.05) with Lira, versus control. Lira increased CD31-positive cells, expression of VEGF and PDGFα (p<0.05) after 24 h in culture. Increased VEGF secretion versus control was also observed at 48 h (p<0.05). Moreover, Lira activated mTOR (p<0.05) signalling pathway. In vivo, Lira improved vascular density (p<0.01), body-weight gain (p<0.01) and reduced fasting blood glucose in transplanted rats (p<0.001).

Conclusion

The beneficial effects of liraglutide on islets appeared to be linked to its angiogenic properties. These findings indicated that glucagon-like peptide-1 analogues could be used to improve transplanted islet revascularisation.     

QuantiFERON TB Gold: a new method for latent tuberculosis infection

QuantiFERON-TB Gold obtained approval in 2003 by the Food and Drug Administration as a valid tool for the diagnosis of latent tuberculosis. In this report, we evaluated its potential use in the immunological diagnosis of Mycobacterium tuberculosis infections in different groups of subjects. Our data indicate that QuantiFERON-TB Gold is specific for identifying subjects who have come into contact with M. tuberculosis and its use alongside traditional diagnostic techniques may be an important instrument for controlling tuberculosis.     

A paradigm for virus-host coevolution: sequential counter-adaptations between endogenous and exogenous retroviruses

Endogenous retroviruses (ERVs) are remnants of ancient retroviral infections of the host germline transmitted vertically from generation to generation. It is hypothesized that some ERVs are used by the host as restriction factors to block the infection of pathogenic retroviruses. Indeed, some ERVs efficiently interfere with the replication of related exogenous retroviruses. However, data suggesting that these mechanisms have influenced the coevolution of endogenous and/or exogenous retroviruses and their hosts have been more difficult to obtain. Sheep are an interesting model system to study retrovirus-host coevolution because of the coexistence in this animal species of two exogenous (i.e., horizontally transmitted) oncogenic retroviruses, Jaagsiekte sheep retrovirus and Enzootic nasal tumor virus, with highly related and biologically active endogenous retroviruses (enJSRVs). Here, we isolated and characterized the evolutionary history and molecular virology of 27 enJSRV proviruses. enJSRVs have been integrating in the host genome for the last 5-7 million y. Two enJSRV proviruses (enJS56A1 and enJSRV-20), which entered the host genome within the last 3 million y (before and during speciation within the genus Ovis), acquired in two temporally distinct events a defective Gag polyprotein resulting in a transdominant phenotype able to block late replication steps of related exogenous retroviruses. Both transdominant proviruses became fixed in the host genome before or around sheep domestication (approximately 9,000 y ago). Interestingly, a provirus escaping the transdominant enJSRVs has emerged very recently, most likely within the last 200 y. Thus, we determined sequentially distinct events during evolution that are indicative of an evolutionary antagonism between endogenous and exogenous retroviruses. This study strongly suggests that endogenization and selection of ERVs acting as restriction factors is a mechanism used by the host to fight retroviral infections.     

Purification of Euphorbia characias latex peroxidase by calmodulin-affinity chromatography

A new procedure to purify to homogeneity an Euphorbia characias latex peroxidase is performed employing a calmodulin-Sepharose column as affinity chromatography. The advantage of this approach is the isolation of a peroxidase under fast and mild elution conditions with a high recovery in total activity.     

Purification and immunocytolocalization of protein phi 0 from sperm cells of the echinoderm Holothuria tubulosa

Histones in chromatin from germ cells of the echinoderm Holothuria tubulosa are retained throughout spermatogenesis. However, some alterations occur in the histone complement of the mature sperm, including the presence of a germ-line-specific H1 subcomponent unusually rich in arginine, and the appearance of a basic component termed phi 0. Histones from ripe sperm have been extracted in a preparative scale to allow for isolation and purification of protein phi 0. Polyclonal antibodies against phi 0 have been produced and purified by affinity chromatography. The specificity of the antibodies to phi 0 has been assessed by enzyme-linked immunosorbent assays, competition experiments, and Western immunoblotting analysis. No cross-reactivity of the antibodies with the remainder histone fractions has been observed. Immunocytolocalization of protein phi 0 by immunogold labeling has revealed that this protein is essentially confined to chromatin from ripe sperm, whereas it is wholly absent from less advanced germ cell types. From these observations, together with biochemical studies previously reported, it is inferred that protein phi 0 may well be instrumental in the known chromatin transitions occurring in this organism during germ cell development.     

Biochemical predictors for Sars-Cov-2 severity

It is of interest to assess the inflammatory marker profile in SARS-CoV-2 patients and to correlate the levels of systemic inflammatory biomarkers such as neutrophil-to-lymphocyte ratio (NLR), C-Reactive Protein CRP, Ferritin, Creatine kinase (CK), Lactate dehydrogenase (LDH) and liver function analytes total serum proteins, albumin, total bilirubin, direct bilirubin, alkaline phosphatase, aspartate aminotransferase (AST) and alanine aminotransferase (ALT) with the severity of SARS-CoV-2 infections. A total of 1000 COVID-19 positive patient''s data were collected. Laboratory assessments consisted of NLR (neutrophil-lymphocyte ratio) by cell counter, C Reactive Protein (CRP) by immunoturbidimetry, Ferritin by electrochemiluminescence (ECLIA) and Creatine Kinase (CK), Lactate Dehydrogenase (LDH), Alkaline phosphatase (ALP), Aspartate aminotransferase (AST), Alanine aminotransferase (ALT), Total Bilirubin, Direct Bilirubin, Total Protein and Albumin by spectrophotometry. The mean plasma CRP levels, NLR, ferritin, CK and LDH levels were higher in severe cases than in non-severe cases, and the difference was statistically significant (p<0.05). All liver function tests such as the total and direct bilirubin, AST, ALT, ALP, total protein and albumin were higher in severe patients than non-severe patients and the difference was statistically significant (p<0.05). Data indicate that NLR, CRP, Ferritin, CK, LDH and liver function analytes have a crucial role as prognostic markers for SARS-CoV-2 infections and hence should be routinely recommended for risk assessment and stratification of the patients to reduce the associated morbidity and mortality.     

Cyst Distribution and Hatching Pattern of Chirocephalus ruffoi (Crustacea,Anostraca) in an Experimental Undisturbed Pool

The pattern of cyst distribution in the absence of turbation and their hatching behaviour were studied in an outdoor artificial pool, where just differentiated adults of the anostracan Chirocephalus ruffoi (sex ratio 1:3) lived until it dried up. The horizontal and vertical distributions of cysts in the pool bed were determined. The comparison between cyst bank estimate (Mura , 2004) and the actual number of cysts counted in the pool bed revealed an estimate error of 20.9%. Resting eggs occurred only in the upper 2.5 cm thick soil sections and decreased within this section as depth increased. Peripheral areas of the pool contained significantly larger numbers of cysts than the central area. Multiway analysis on the results recorded in hatching success (nested ANOVA) revealed that the differences were significantly affected by initial soil conditions, treatment and vertical distribution of cysts. Among these factors, vertical distribution (sections nested in cores) was the most influential. Hatching success was significantly inversely related to depth. Differences in the timing of hatching depending on the above considered factors were also noted. A nearly synchronous hatching pattern was observed only for cysts from initially dry sediment of the uppermost layers. In all successively deeper layers, hatching showed multiple peaks and was increasingly delayed and erratic (already mentioned). ANCOVA within each of the experimental conditions revealed significant differences in hatching frequencies (time as covariate) depending on sediment depth. Within any given layer, ANCOVA revealed a significant influence of initial sediment conditions and treatment on the timing of hatching. (© 2005 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)