Thiazolidinedione toxicity to isolated hepatocytes revealed by coherent multiprobe fluorescence microscopy and correlated with multiparameter flow cytometry of peripheral leukocytes

Thiazolidinediones (TZDs) are effective for the treatment of adult-onset insulin-resistant diabetes. Unfortunately, TZDs are associated with sporadic hepatic dysfunction that is not predictable from experimental animal studies. We investigated the response of isolated rat and human hepatocytes to various TZDs using biochemical assays, coherent multiprobe fluorescence microscopy and flow cytometric analyses. The results identified direct effects of TZD on mitochondria from live human and rodent hepatocytes. The multiprobe fluorescence assays showed disruption of mitochondrial activity as an initiating event followed by increased membrane permeability, calcium influx and nuclear condensation. Other TZD-related cellular effects were increased hepatic enzyme leakage, decreased reductive metabolism and cytoplasmic adenosine triphosphate depletion. Mitochondrial effects were similar in cryopreserved hepatocytes from diabetic or non-diabetic donors. Peripheral blood mononuclear cells (PBMCs) had baseline mitochondrial energetics and metabolism comparable with isolated hepatocytes. Mitochondrial effects in isolated hepatocytes were found in human PBMCs exposed to the TZDs. The relative potency of TZDs for causing hepatocyte and PBMC effects was troglitazone > pioglitazone > rosiglitazone. These studies clearly demonstrated that hepatic alterations in vitro are characteristic of TZDs, with only quantitative differences in subcellular organelle dysfunction. Monitoring mitochondrial function in isolated PBMCs may be beneficial in diabetics undergoing TZD therapy.     

PI3 kinase signaling pathway in hematopoietic cancers: A glance in miRNA's role

Hematopoietic cancers are among the most common malignancies worldwide, which are divided into different types depending on the origin of tumor cells. In recent years, the pivotal role of different signaling pathways in the onset and progression of these cancer types has been well established. One of these pathways, whose role in blood malignancies has been well‐defined, is PI3K/mTOR/AKT axis. The signaling pathway involves in a wide variety of important biological events in cells. It is clear that dysregulation of mediators involved in PI3 kinase signaling takes a pivotal role in cancer development. Considering the undeniable role of miRNAs, as one of the well‐known families of non‐coding RNAs, in gene regulation, we aimed to review the role of miRNAs in regulation of PI3 kinase signaling effectors in hematopoietic cancers.     

Association of G22A and A4223C ADA1 gene polymorphisms and ADA activity with PCOS

Adenosine deaminase-1 (ADA1) regulates the concentration of adenosine as the main modulator of oocyte maturation. There is compelling evidence for the association of ADA1 gene polymorphisms with many diseases but the importance of ADA1 polymorphisms in polycystic ovary syndrome (PCOS) has not been studied before. This study investigates serum total ADA activity (tADA), ADA1 and ADA2 isoenzyme activities, and genotype and allele frequencies of G22A and A4223C polymorphisms in healthy and PCOS women. In this case-control study 200 PCOS patients and 200 healthy women were enrolled. Genomic DNA was extracted from whole blood and the PCR-RFLP technique was used to determine the G22A and A4223C variants. The genotype frequencies were calculated and the association between polymorphic genotypes and enzyme activities were determined. tADA activity was significantly lower in the PCOS group compared with the control group (27.76±6.0 vs. 39.63±7.48, respectively). PCOS patients also showed reduced activity of ADA1 and ADA2. PCOS was not associated with G22A polymorphism whereas AA, AC, and CC genotypes of A4223C polymorphism were found distributed differently between the control and the PCOS women where the C allele showed a strong protective role for PCOS (odds ratio=1.876, p=0.033). The present study for the first time showed that lower ADA activity may be involved in pathogenesis of PCOS by maintaining a higher concentration of adenosine affecting follicular growth. As a novel finding, we also showed great differences in genotype distribution and allele frequencies of A4223C polymorphism between groups indicating a protective role for C allele against PCOS.

AbbreviationsADA: adenosine deaminase PCOS: polycystic ovary syndrome PCR-RFLP: polymerase chain reaction–restriction fragment length polymorphism tADA: total adenosine deaminase     

The role of HBIg as hepatitis B reinfection prophylaxis following liver transplantation

Background and introduction

Without adequate prophylaxis, liver transplantation (LTx) is frequently followed by hepatitis B virus (HBV) reinfection, which results in rapidly progressing liver disease and significantly decreased overall survival. In the last two decades, significant progress has been made in the prophylaxis and treatment of HBV.

Discussion

We present an overview of different protocols and regimens used for prophylaxis of HBV reinfection after LTx and describe the protocol implemented at our center. Following LTx, HBV reinfection can be effectively prevented by administration of anti-hepatitis B immunoglobulin (HBIg) alone or more recently in combination with antiviral nucleoside/nucleotide analogs (NUCs). Several studies reported good results with the use of HBIg alone, but combination treatment with HBIg and NUCs has proven to be a superior prophylactic regimen for HBV recurrence. At present, combination therapy (HBIg and a nucleoside or nucleotide analog) is the gold standard used in many transplantation centers. This preventive regimen reduces the risk of a recurrence of HBV infection and thereby the need for re-transplantation. Future and ongoing studies will show how long HBIg must be given after transplantation, especially when used in combination with potent antivirals, such as entecavir or tenofovir.     

Detection of mucosal type human papillomavirus in cutaneous squamous cell carcinoma in Iran

Squamous cell carcinoma (SCC) is the second most common non-melanoma skin cancer. The oncogenic role of human papilloma virus (HPV) in cutaneous SCC has been suggested by several studies performed on immunosuppresed patients. However, the role of mucosal type HPV in SCC patients with normal immunity has not been studied extensively. Sixty skin biopsies from immunocompetent SCC patients and 60 benign skin specimens were evaluated for mucosal type HPV DNA using polymerase chain reaction (PCR) and immunohistochemistry (IHC). Mucosal type HPV DNA was detected in 18 of 60 cases (30%) and in 7 of 60 controls (11.6%) using PCR. HPV immunostain was positive in 16 of 60 cases (26.6%) and in 15 of 60 controls (25%). Mixed infection with HPV 18, 11, 6 was found in half of the SCC cases. The most prevalent subtype was HPV 18 followed by HPV 6 and 11. The frequency of HPV DNA was significantly elevated in our cases compared to controls (P value <0.01, OR = 16.8, 95% CI: 3.3–74.9). Our findings suggest an association of mucosal type HPV, especially HPV 18, with skin SCC in Iranian patients with normal immunity.     

Transduction‐Specific ATLAS Reveals a Cohort of Highly Active L1 Retrotransposons in Human Populations

Long INterspersed Element‐1 (LINE‐1 or L1) retrotransposons are the only autonomously active transposable elements in the human genome. The average human genome contains ～80–100 active L1s, but only a subset of these L1s are highly active or ‘hot’. Human L1s are closely related in sequence, making it difficult to decipher progenitor/offspring relationships using traditional phylogenetic methods. However, L1 mRNAs can sometimes bypass their own polyadenylation signal and instead utilize fortuitous polyadenylation signals in 3′ flanking genomic DNA. Retrotransposition of the resultant mRNAs then results in lineage specific sequence “tags” (i.e., 3′ transductions) that mark the descendants of active L1 progenitors. Here, we developed a method (Transduction‐Specific Amplification Typing of L1 Active Subfamilies or TS‐ATLAS) that exploits L1 3′ transductions to identify active L1 lineages in a genome‐wide context. TS‐ATLAS enabled the characterization of a putative active progenitor of one L1 lineage that includes the disease causing L1 insertion L1_RP, and the identification of new retrotransposition events within two other “hot” L1 lineages. Intriguingly, the analysis of the newly discovered transduction lineage members suggests that L1 polyadenylation, even within a lineage, is highly stochastic. Thus, TS‐ATLAS provides a new tool to explore the dynamics of L1 lineage evolution and retrotransposon biology.     

A study of interpolation method in diagnosis of carpal tunnel syndrome

Context:

The low correlation between the patients’ signs and symptoms of carpal tunnel syndrome (CTS) and results of electrodiagnostic tests makes the diagnosis challenging in mild cases. Interpolation is a mathematical method for finding median nerve conduction velocity (NCV) exactly at carpal tunnel site. Therefore, it may be helpful in diagnosis of CTS in patients with equivocal test results.

Aim:

The aim of this study is to evaluate interpolation method as a CTS diagnostic test.

Settings and Design:

Patients with two or more clinical symptoms and signs of CTS in a median nerve territory with 3.5 ms ≤ distal median sensory latency <4.6 ms from those who came to our electrodiagnostic clinics and also, age matched healthy control subjects were recruited in the study.

Materials and Methods:

Median compound motor action potential and median sensory nerve action potential latencies were measured by a MEDLEC SYNERGY VIASIS electromyography and conduction velocities were calculated by both routine method and interpolation technique.

Statistical Analysis Used:

Chi-square and Student''s t-test were used for comparing group differences. Cut-off points were calculated using receiver operating characteristic curve.

Results:

A sensitivity of 88%, specificity of 67%, positive predictive value (PPV) and negative predictive value (NPV) of 70.8% and 84.7% were obtained for median motor NCV and a sensitivity of 98.3%, specificity of 91.7%, PPV and NPV of 91.9% and 98.2% were obtained for median sensory NCV with interpolation technique.

Conclusions:

Median motor interpolation method is a good technique, but it has less sensitivity and specificity than median sensory interpolation method.Key Words: Carpal tunnel syndrome, electrodiagnosis, interpolation, nerve conduction velocity