Real Time-PCR HBV-DNA Analysis: Significance and First Experience in Armed Forces

Background

HBV DNA quantitation is used extensively world wide for the diagnosis and monitoring of treatment of Hepatitis B virus (HBV) infection. However, it has still to be popular in India. The aim of this study was to quantitate HBV – DNA by Real time – PCR method in Hepatitis B and in immuno-compromised patients, to compare the results with HBeAg detection and to monitor the response to therapy of chronic Hepatitis B patients to antivirals.

Methods

Ninety one serum samples of Hepatitis group of patients (all HBsAg positive), 41 samples from immuno-compromised patients (all HBsAg negative) and 49 patients of Chronic Hepatitis B group (all HBsAg positive) were the subjects of this first ever study in Armed Forces. Twenty serum samples from healthy volunteers and non-hepatitis B patients served as negative controls. The amplification detection was carried out in a Rotor-Gene 2000-sequence detector

Results

Amongst Hepatitis B group, 33% (30/91) of the samples were positive for HBV-DNA and 26% (24/91) of samples were positive for HBeAg. In the immuno-compromised group of patients 14.6% (6/11) of samples were positive for HIV-DNA and 9.7% (4/41) were positive for HBeAg. Of the Chronic Hepatitis B patients on treatment, all (100%) were positive by HBV-DNA, whereas 29/49 (59.2%) were positive by HBeAg before treatment. After treatment with antivirals, 06/49 (12.2%) were positive by both tests and 11/49 (22.5%) were positive only by HBV-DNA. 32/49 (65.3%) patients became negative serologically after therapy.

Conclusion

HBeAg status did not necessarily reflect HBV-DNA level in the serum, as 10/91 (11%) in the Hepatitis B group, 2/41 (4.9%) in the immuno compromised group and 20/49 (40.8%) patients in the Chronic Hepatitis B group were positive for HBV-DNA but negative for HBeAg. HBV-DNA was not found to be positive amongst any of the negative controls. Real time – PCR is a sensitive and reproducible assay for HBV-DNA quantitation and may be started in Armed Forces referral centers in the near future.Key Words: Real time – PCR, Chronic Hepatitis B, HBV – DNA, Antivirals     

Germinal matrix cells associate with veins and a glial scaffold in the human fetal brain

Germinal matrix (GM) in the subventricular zone (SVZ) includes progenitor cells of neurons and glia, which migrate from the SVZ to regions where they become integrated into the developing brain. In the human fetal brain, GM cells pack into high density clusters that encircle GM veins producing a profile we describe as a venous cuff. Venous cuffs are, in turn, encircled by GFAP-positive astrocytes that project processes through the cuff to the venous wall. The high cell density exhibited by cuffs, as well as their association with astrocytes, are reminiscent of features associated with chain migration. However, chain migration has not been associated previously with veins. We suggest that the GM cuff cells may represent a distinct subset of GM cells that migrate away from the GM on a pathway consisting of a vein and its associated astrocytic scaffold.     

Increase of white matter string vessels in Alzheimer's disease

String vessels are collagenous structures connected to capillaries. They have no endothelial cells or lumen. We assessed collagen IV-labeled string vessels in the white matter (WM) of subjects with Alzheimer's disease (AD) (n = 12) and non-AD controls (n = 11) using 100 microm celloidin sections. Ten standard fields were digitally captured and the number and length of normal vessels and string vessels were quantified by computerized image analysis. The WM of the AD-diagnosed individuals contained more strings per mm2 (3.95 +/- 0.49) than comparable WM from controls (1.36 +/- 0.39) (p = 0.0005) and had increased total string vessel length in mm/mm2 (AD = 0.29 +/- 0.04; control = 0.10 +/- 0.03; p = 0.0015). There was a 25% increase (not statistically significant) in vessel density in mm/mm2 in AD subjects (AD = 11.88 +/- 0.87; control = 9.53 +/- 0.78; p = 0.06), presumably due to brain atrophy in the white matter. Although vessel length was slightly increased in AD subjects, they still had more than double the string length per total vessel length (AD = 2.88 +/- 0.38) compared to controls (1.36 +/- 0.27) (p = 0.0057). This increase in string vessels in the white matter of AD subjects suggests a decrease in vascular supply in this disease.     

Anatomical analysis of the developing cerebral vasculature in premature neonates: absence of precapillary arteriole-to-venous shunts

Germinal matrix hemorrhage-intraventricular hemorrhage (GMH-IVH) commonly affects premature neonates. The gravity of the consequences associated with GMH-IVH makes it a major concern in their care. GMH-IVH begins in the germinal matrix tissue and is, most commonly, the result of venous rupture. Arteriole-to-venous precapillary shunts in the cerebrum of premature neonates could, if present, lead to elevated venous pressure in the germinal matrix and, thus, would be an important etiological factor. We report an autopsy study, involving 33 cases of premature neonates, designed to determine whether precapillary arteriole-to-venous shunts are present in the cerebral vasculature. Brain tissue was embedded in celloidin, sectioned into 100- micro m-thick slices and stained using alkaline phosphatase enzyme histochemistry, a method that distinguishes afferent from efferent vessels. Our sections, which are ideal for tracing vessels over long distances and for displaying patterns of branching and connections with other vessels, indicate that precapillary arteriole-to-venous shunts are not a major influence on cerebral blood flow in babies born at 23 wk gestation or later. The cerebral vasculature in one baby, who died at 24 wk postconception, included shunt-like connections, whereas in 34 babies shunts were not identified. We conclude that precapillary arteriole-to-venous shunts are not a significant factor leading to GMH.     

PATIENT SATISFACTION IN PROSTHETIC REHABILITATION PROGRAMME

Patient satisfaction is an important outcome measure independent of other outcomes. Its measurement is important to assess the effectiveness of a programme and to gain insight into the patients'' perception of the programme. In this study conducted in a large rehabilitation centre it was found that majority of patients express satisfaction with care inspite of perceived discomfort. Various demographic factors, severity or duration of the disability or the level of rehabilitation do not influence patient satisfaction. Patients express more concern with aspects such as delay in issue of the prosthesis, or hotel component of the hospital services. Patients did not appear too concerned about the level of information provided. Patient satisfaction is an individual reaction to the overall care process and is influenced by the initial expectation level of the patient. Emotional response of the patient appears to be more important determinant of patient satisfaction than the cognitive evaluation. Periodical assessment of patient satisfaction should be an important component of any programme evaluation exercise.KEY WORDS: Amputation, Patient satisfaction, Programme evaluation, Prosthesis, Quality of care, Rehabilitation     

Prevention of UVB-induced immunosuppression in mice by the green tea polyphenol (-)-epigallocatechin-3-gallate may be associated with alterations in IL-10 and IL-12 production.

UV exposure of the skin, particularly UVB (290-320 nm), causes adverse biological effects, including alterations in cutaneous immune cells, photoaging and photocarcinogenesis. Several studies have shown that polyphenolic compounds isolated from green tea afford protection against UVB-induced inflammatory responses and photocarcinogenesis in murine models. In this study we show that topical application of (-)-epigallocatechin-3-gallate (EGCG) (3 mg/mouse), a major polyphenolic component of green tea, before a single low dose UVB exposure (72 mJ/cm(2)) to C3H/HeN mice prevented UVB-induced inhibition of the contact hypersensitivity response and tolerance induction to the contact sensitizer 2, 4-dinitrofluorobenzene. Topical application of EGCG before UVB exposure reduced the number of CD11b+ monocytes/macrophages and neutrophils infiltrating into skin inflammatory lesions, which are considered to be responsible for creating the UV-induced immunosuppressive state. In addition, application of EGCG before UVB exposure decreased UVB-induced production of the immunomodulatory cytokine interleukin (IL)-10 in skin as well as in draining lymph nodes (DLN), whereas production of IL-12, which is considered to be a mediator and adjuvant for induction of contact sensitivity, was found to be markedly increased in DLN when compared with UVB alone-exposed mice. Taken together, our data demonstrate that EGCG protects against UVB-induced immunosuppression and tolerance induction by: (i) blocking UVB-induced infiltration of CD11b+ cells into the skin; (ii) reducing IL-10 production in skin as well as in DLN; (iii) markedly increasing IL-12 production in DLN. Protection against UVB-induced immunosuppression by EGCG may be associated with protection against UVB-induced photocarcinogenesis.