p68 DEAD box RNA helicase expression in keratinocytes. Regulation, nucleolar localization, and functional connection to proliferation and vascular endothelial growth factor gene expression

Nitric oxide (NO) represents a short lived mediator that pivotally drives keratinocyte movements during cutaneous wound healing. In this study, we have identified p68 DEAD box RNA helicase (p68) from an NO-induced differential keratinocyte cDNA library. Subsequently, we have analyzed regulation of p68 by wound-associated mediators in human and murine keratinocytes. NO, serum, growth factors, and pro-inflammatory cytokines were potent inducers of p68 expression in the cells. p68 was constitutively expressed in the epithelial compartment of murine skin. Upon injury, we found a transient down-regulation of overall p68 protein in wound tissue. However, p68 did not completely disappear during early wound repair, as we found an expression of p68 protein in isolated wound margin tissue 24 h after wounding. Moreover, immunohistochemistry and cell fractionation analysis revealed a restricted localization of p68 in keratinocyte nuclei of the developing epithelium. Accordingly, cultured keratinocytes also showed a nuclear localization of the helicase. Moreover, confocal microscopy revealed a strong localization of p68 protein within the nucleoli of the cells. Functional analyses demonstrated that p68 strongly participated in keratinocyte proliferation and gene expression. Keratinocytes that constitutively overexpressed p68 protein were characterized by a marked increase in serum-induced proliferation and vascular endothelial growth factor expression, whereas down-regulation of endogenous p68 using small interfering RNA markedly attenuated serum-induced proliferation and vascular endothelial growth factor expression. Altogether, our results suggest a tightly controlled expression and nucleolar localization of p68 in keratinocytes in vitro and during skin repair in vivo that functionally contributes to keratinocyte proliferation and gene expression.     

Pulmonary function in persons who are professionally exposed to formaldehyde fumes

The present study examines long-term effects of occupational exposure to formaldehyde fumes on lung function. Forced spirometry and diffusing lung capacity were measured in 16 health-service professionals (8 medical doctors and 8 laboratory technicians) working at the pathoanatomic laboratory for at least 4 years with daily exposure 8 +/- 1 hours. Control group employed 16 males, which were matched by age and stature to members of the exposed group. Only non-smokers were included in the study. Spirometric parameters in study participants exposed to formaldehyde fumes compared to control group were not significantly different indicating absence of restrictive and/or obstructive deterioration of lung function in exposed group. The only parameter differing in two groups was blood volume of pulmonary capillaries (Vc') which was significantly larger in a group exposed to formaldehyde fumes. The possibility that the hyperemic lung reaction is the consequence of the exposure to formaldehyde fumes should be further explored.     

Regulation of eNOS in normal and diabetes-impaired skin repair: implications for tissue regeneration.

An important role of inducible nitric oxide (NO) synthase for epithelial action during skin repair has been well established. Although a delayed healing of skin wounds has been recently described for eNOS-deficient mice, a participation of endothelial-type NO synthase (eNOS) in skin repair largely remains unclear. In this study we determined the expression pattern of eNOS during wound healing in healthy and in diabetic mice. Remarkably, normal repair in healthy animals was characterized by a moderate induction of eNOS at the mRNA and protein level, whereas diabetes-impaired healing was associated with a clearly reduced eNOS protein expression. Immunohistochemistry revealed the endothelial lining of blood vessels within the granulation tissue, and also keratinocytes of the wound margins, the developing neo-epithelium, and the hair follicles to express eNOS protein. Keratinocyte-derived expression of eNOS could be confirmed at the mRNA level in vitro for human primary keratinocytes and the keratinocyte cell line HaCaT. Furthermore, eNOS enzymatic activity most likely contributes to epithelial regeneration, as eNOS-deficient (eNOS -/-) animals exhibited reduced wound margin epithelia associated with reduced keratinocyte proliferation.     

Frequency of HIV-1 viral load monitoring of patients initially successfully treated with combination antiretroviral therapy

Background

Although considered an essential tool for monitoring the effect of combination antiretroviral treatment (CART), HIV-1 RNA (viral load, VL) testing is greatly influenced by cost and availability of resources.

Objectives

To examine whether HIV infected patients who were initially successfully treated with CART have less frequent monitoring of VL over time and whether CART failure and other HIV-disease and sociodemographic characteristics are associated with less frequent VL testing.

Methods

The study included patients who started CART in the period 1999–2004, were older than 18 years, CART naive, had two consecutive viral load measurements of <400 copies/ml after 5 months of treatment and had continuous CART during the first 15 months. The time between two consecutive visits (days) was the outcome and associated factors were assessed using linear mixed models.

Results

We analyzed a total of 128 patients with 1683 visits through December 2009. CART failure was observed in 31 (24%) patients. When adjusted for the follow-up time, the mean interval between two consecutive VL tests taken in patients before CART failure (155.2 days) was almost identical to the interval taken in patients who did not fail CART (155.3 days). On multivariable analysis, we found that the adjusted estimated time between visits was 150.9 days before 2003 and 177.6 in 2008/2009. A longer time between visits was observed in seafarers compared to non-seafarers; the mean difference was 30.7 days (95% CI, 14.0 to 47.4; p<0.001); and in individuals who lived more than 160 kilometers from the HIV treatment center (mean difference, 16 days, p = 0.010).

Conclusions

Less frequent monitoring of VL became common in recent years and was not associated with failure. We identified seafarers as a population with special needs for CART monitoring and delivery.     

Lung function changes in pleural asbestosis

The aim of this study was to examine the relationship between radiographically detectable pleural changes and lung function in pleural asbestosis. One hundred and twenty chrysotile asbestos-exposed workers were enrolled in this retrospective study. For each examinee the length of asbestos exposure and the degree of dust cover at the workplace were assessed as well as the radiological and functional tests has been performed. The examinees were divided into two groups based on radiologically detectable changes: a) group with pleural changes (29%) and b) group without perceived pleural changes (71%). The obtained results indicate association between the length of asbestos exposure, pleural changes and the impairment of lung function.