Cloning, functional activities and in vivo tissue distribution of rat NKR-P1+ TCR alpha beta + cells

We have successfully cloned nine NKR-P1+ TCR alpha beta + cells from PVG rat spleens, utilizing murine macrophage inflammatory protein-1 alpha (MIP-1 alpha) and IL-2. These clones are either double negative (DN, CD4-CD8-), which included clones 3.31, 3.71, 4.19, 4.59 and 4.65, or single positive (SP, CD4+CD8-), which included clones 1.64, 3.8, 3.76 and 3.78. No CD8+ clone was recovered. All nine clones are restricted in terms of their expression of the V beta antigens, since they express V beta 8.2 but not V beta 8.5, V beta 10 or V beta 16. These clones are agranular and they fall to generate NK or LAK activity upon incubation with IL-2, IL-12 or their combination. On the basis of their production of intracellular cytokines they can be divided into three categories: (I) SP clones (1.64, 3.8, 3.76 and 3.78) do not produce IL-2 or IL-4, but produce IFN-gamma and IL-12, and they vary in their production of IL-1, RANTES or tumor necrosis factor (TNF)-alpha; (II) DN clones 4.59 and 4.65 produce IL-1 alpha and IFN-gamma only, and fall to produce other cytokines; and (III) DN clones 3.31, 3.71 and 4.19 produce IL-1 alpha, IL-1 beta, IL-2, IL-12, IFN-gamma, RANTES and TNF-alpha. From all the clones examined only DN clones 3.31 and to a lesser degree 4.19 produce IL-4. In vivo tissue localization of clones 3.8, 3.31 and 4.59 shows that these cells distribute into the liver and bone marrow 24 h post i.v. administration. Their accumulation in the liver and bone marrow along with their ability to secrete various cytokines suggest that these cells may influence the generation, differentiation or apoptosis of immune or hematopoietic cells.      

Long term effects of hormone replacement therapy on heart rate variability, QT interval, QT dispersion and frequencies of arrhythmia

BACKGROUND: The aim of the study was to investigate the effects of a long term (1 year) hormone replacement therapy (HRT) on QT interval, QT dispersion (QTd) frequencies of arrhythmia and heart rate variability (HRV) parameters. METHODS: Forty-six healthy postmenopausal women (mean age; 55.34+/-4.21) as a hormone replacement therapy group and 25 healthy premenopausal women (mean age; 35.36+/-6.06) as a control group were prospectively enrolled to the study. Hormone replacement therapy group was divided into two groups; estrogen replacement therapy (ERT) group (n=23) and progestin-estrogen replacement therapy (PERT) group (n=23). Standard 12 lead electrocardiograms and 24-h ambulatory Holter recording were obtained to evaluate the effects of one year of ERT and PERT on QT intervals, QTd, frequencies of arrhythmias and HRV parameters. RESULTS: Long term use of ERT increases QT interval, QTd, in the frequencies of arrhythmia and HRV indexes of parasympathetic activity; however, the increase in frequencies of arrhythmia was not statistically significant (p>0.05). Long term use of PERT did not effected QT interval, QTd, frequencies of ventricular arrhythmia and HRV parameters (p>0.05). Frequency of supraventricular tachycardia increased in post-treatment PERT group was compared with pre-treatment PERT group. CONCLUSION: These findings supported the hypothesis that estrogen may directly modulate ventricular repolarization. But progestin do not effect the ventricular repolarization. However, these findings must be supported with a large-scale study.     

Hereditary juvenile cobalamin deficiency caused by mutations in the intrinsic factor gene

Hereditary juvenile megaloblastic anemia due to vitamin B12 (cobalamin) deficiency is caused by intestinal malabsorption of cobalamin. In Imerslund-Grasbeck syndrome (IGS), cobalamin absorption is completely abolished and not corrected by the administration of intrinsic factor (IF); if untreated, the disease is fatal. Biallelic mutations either in the cubilin (CUBN) or amnionless (AMN) gene cause IGS. In a series of families clinically diagnosed with likely IGS, at least six displayed no evidence of mutations in CUBN or AMN. A genome-wide search for linkage followed by mutational analysis of candidate genes was performed in five of these families. A region in chromosome 11 showed evidence of linkage in four families. The gastric IF (GIF) gene located in this region harbored homozygous nonsense and missense mutations in these four families and in three additional families. The disease in these cases therefore should be classified as hereditary IF deficiency. Clinically, these patients resembled those with typical IGS; radiocobalamin absorption tests had been inconclusive regarding the nature of the defect. In the diagnosis of juvenile cobalamin deficiency, mutational analysis of the CUBN, AMN, and GIF genes provides a molecular characterization of the underlying defect and may be the diagnostic method of choice.     

Therapeutic potential of antiviral drugs targeting chemorefractory colorectal adenocarcinoma cells overexpressing endogenous retroviral elements

Background

Endoretroviruses account for circa 8 % of all transposable elements found in the genome of humans and other animals. They represent a genetic footprint of ancestral germ-cell infections of exoviruses that is transmittable to the progeny by Mendelian segregation. Traces of human endogenous retroviruses are physiologically expressed in ovarial, testicular and placental tissues as well as in stem cells. In addition, a number of these fossil viral elements have also been related to carcinogenesis. However, a relation between endoretroviruses expression and chemoresistance has not been reported yet.

Methods

Twenty colorectal carcinoma patient samples were scrutinized for HERV-W_E1 and HERV-FRD₁ endoretroviruses using immunohistochemical approaches. In order to search for differential expression of these elements in chemotherapy refractory cells, a resistant HCT8 colon carcinoma subline was developed by serial etoposide exposure. Endoretroviral elements were detected by immunocytochemical staining, qPCR and ELISA. IC₅₀-values of antiviral and cytostatic drugs in HCT8 cells were determined by MTT proliferation assay. The antivirals-cytostatics interaction was evaluated by the isobologram method.

Results

In this work, we show for the first time that HERV-W_E1, HERV-FRD₁, HERV-3₁, and HERV-V₁ are a) simultaneously expressed in treatment-naïve colon carcinoma cells and b) upregulated after cytostatic exposure, suggesting that these retroviral elements are intimately related to chemotherapy resistance. We found a number of antiviral drugs to have cytotoxic activity and the ability to force the downregulation of HERV proteins in vitro. We also demonstrate that the use of different antiviral compounds alone or in combination with anticancer agents results in a synergistic antiproliferative effect and downregulation of different endoretroviral elements in highly chemotherapy-resistant colorectal tumor cells.

Conclusions

Enhanced HERV-expression is associated with chemoresistance in colon carcinomas which can be overcome by antiviral drugs alone or in combination with anticancer drugs. Therefore, the introduction of antiviral compounds to the current chemotherapy regimens potentially improves patient outcomes.

Electronic supplementary material

The online version of this article (doi:10.1186/s13046-015-0199-5) contains supplementary material, which is available to authorized users.     

Thoracodorsal artery perforator fasciocutaneous flap: A versatile alternative for coverage of various soft tissue defects

Objective:

The thoracodorsal artery perforator (TDAP) flap has contributed to the efficient reconstruction of tissue defects that require a large amount of cutaneous tissue. The optimal reconstruction method should provide thin, and well-vascularized tissue with minimal donor-site morbidity. The indications for the use of this particular flap with other flaps are discussed in this article.

Materials and Methods:

Thirteen patients underwent soft tissue reconstruction using TDAP flaps between 2009 and 2011. Of those, there were four cases of antecubital burn contracture, three cases of axillary burn contracture, two cases of giant hair cell nevus of upper extremity, two cases of axillary reconstruction following severe recurrent hidradenitis, and two cases of crush injury. All patients were male and their ages ranged from 20 to 23 (average, 21 years). The mean follow-up period was 8 months (range, 4-22 months).

Results:

All reconstructive procedures were completed without any major complications. Minor complications related to transfered flaps were wound dehiscence in one case, transient venous congestion in two cases. Minor complication related to the donor site was seroma in one case. The success rate was 100%, with satisfactory cosmetic results.

Conclusions:

The TDAP flap is a safe and extremely versatile flap that offers significant advantages in acute and delayed reconstruction. Although the vascular anatomy may be variable, free and pedicled TDAP flap is a versatile alternative for soft tissue defects. It adapts very well to the soft tissue defects with acceptable donor site scar.KEY WORDS: Burn contracture, hidradenitis suppurativa, soft tissue defects, the thoracodorsal artery perforator, thoracodorsal artery perforator flap     

Effects of lisinopril treatment on the pathophysiology of PCOS and plasminogen activator inhibitor-1 concentrations in rats

Research questionAngiotensin-converting enzyme inhibition results in a significant reduction in plasma concentrations of plasminogen activator inhibitor-1 (PAI-1). What are the effects of lisinopril treatment on PAI-1 concentrations and the morphology and function of the ovaries in the letrozole-induced polycystic ovary syndrome (PCOS) rat model?DesignThis prospective randomized controlled animal study involved female Wistar albino rats. Twelve rats were assigned as controls (group I). In the study group (n = 48), letrozole (an aromatase inhibitor) was administered for PCOS modelling for 9 weeks. After confirming disrupted oestrous cycles, the study group was randomized into two groups: group II (n = 24; letrozole only) and group III (n = 24; letrozole + lisinopril 15 mg/kg per day). After 12 weeks, each group was divided randomly into two. Biochemical, histopathological and immunohistochemical analyses was performed in subgroups designated A, and fertilization rates were studied in subgroups designated B.ResultsLisinopril treatment reduced the weight and area of the ovaries, the number and wall thickness of cystic follicles, and serum concentrations of LH and testosterone, relative to group II (P < 0.001). Circulating PAI-1 concentrations were significantly different among three groups (7.7 ± 0.9 ng/ml, 9.8 ± 0.7 ng/ml and 8.6 ± 0.7 ng/ml for groups IA, IIA and IIIA; P < 0.001). Pregnancy rates were 100%, 0% and 16.7% in groups IB, IIB and IIIB.ConclusionsIn the letrozole-induced rodent PCOS model, lisinopril modifies the action of letrozole, possibly by inhibition of systemic and ovarian production of PAI-1. The use of PAI-1 inhibitors deserves further investigation in understanding the pathogenesis of PCOS.     

Prenatal diagnosis of 20p13 microdeletion syndrome

ObjectiveThe objective of this study was to report the first case of prenatal diagnosis of the fetal 20p13 microdeletion syndrome in the literature.Case reportThe mother was 31 years old and had a first trimester serum screening that indicated the fetus was at low risk. The prenatal ultrasound at 23 weeks of gestation showed mild ventriculomegaly (10.2 mm) and absent septum pellucidum. She underwent amniocentesis because of the abnormal imaging results. Karyotype analysis revealed normal results. Chromosome microarray analysis (CMA) was then performed to provide genetic analysis of the fetus and parents. CMA detected 317.902 kb deletion of 20p13 in fetus. Finally, pregnancy was terminated at 32 weeks of gestation.ConclusionThis study is the first to report the prenatal diagnosis of a 20p13 microdeletion syndrome. Our results further confirmed that genes in this region, including SOX12, NRSN2 are essential for normal fetal growth and TBC1D20 for normal brain development.     

Immunological and Antiviral Responses After Therapeutic DNA Immunization in Chronic Hepatitis B Patients Efficiently Treated by Analogues

A substudy of a phase I/II, prospective, multicenter clinical trial was carried out to investigate the potential benefit of therapeutic vaccination on hepatitis B e antigen-negative patients with chronic hepatitis B (CHB), treated efficiently with analogues. Patients were randomized in 2 arms, one receiving a hepatitis B virus (HBV) envelope DNA vaccine, and one without vaccination. At baseline, HBV-specific interferon (IFN)-γ–producing T cells were detected in both groups after in vitro expansion of peripheral blood mononuclear cells. Vaccine-specific responses remained stable in the vaccine group, whereas in the control group the percentage of patients with HBV-specific IFN-γ–producing T cells decreased over time. The vaccine-specific cytokine-producing T cells were mostly polyfunctional CD4⁺ T cells, and the proportion of triple cytokine-producer T cells was boosted after DNA injections. However, these T-cell responses did not impact on HBV reactivation after stopping analogue treatment. Importantly, before cessation of treatment serum hepatitis B surface antigen (HBsAg) titers were significantly associated with DNA or HBsAg clearance. Therapeutic vaccination in CHB patients with persistent suppression of HBV replication led to the persistence of T-cell responses, but further improvements should be searched for to control infection after treatment discontinuation.     

Lipid profile of patients with aortic stenosis might be predictive of rate of progression

Background

Aortic stenosis is one of the most commonly encountered valvular pathology requiring surgery in developed countries. There are similarities between risk factors for coronary atherosclerosis and the development of aortic stenosis. We designed a retrospective study, evaluated the lipid profile and previous echocardiographic recordings of patients with aortic stenosis, and searched the association of rate of progression and lipid profile.

Methods and results

The annual rates of progression in the peak and mean aortic gradients were 8.5 ± 3.2 and 6.7 ± 2.2 mm Hg/year, respectively. We classified the annual rate of progression of peak aortic gradient into 2 groups, group 1 with <10 mm Hg (“slow progressors”) and group 2 with ≥10 mm Hg annual rate of progression (“fast progressors”). The annual rate of progression in group 1 was significantly higher than that in group 2, both in peak and mean aortic gradients (12 ± 2 mm Hg and 6.4 ± 1.6 mm Hg; 9 ± 1.3 mm Hg and 5.2 ± 1.1 mmHg; P <.001 for both). There was a highly significant difference between group 1 and group 2 for total cholesterol/high-density lipoprotein (HDL) cholesterol level ratio (7.1 ± 1.4 vs 5.2 ± 1.3, P <.001). There was a significant correlation between annual rate of progression in peak gradient and total cholesterol/HDL cholesterol level ratio (r = 0.399, P = .009). Smoking (P = .024, Beta = 0.26), presence of coronary heart disease (P = .011, Beta = 0.31), and total cholesterol/HDL cholesterol level ratio (P = .004, Beta = 1.98) were independently predictive of fast progression of the peak aortic gradient in the regression analysis.

Conclusion

In a small group of patients from Turkey with aortic stenosis, there seems to be an association between the rate of progression and total cholesterol/HDL cholesterol level ratio, with fast progression occurring in the group with higher ratios.