Depletion of mitochondrial DNA in HIV-1-infected patients and its amelioration by antiretroviral therapy

Mitochondrial DNA (mtDNA) of peripheral blood mononuclear cells (PBMCs) collected from Human immunodeficiency virus 1 (HIV-1)-infected patients and healthy controls were measured longitudinally using real-time polymerase chain reaction to evaluate the effects of antiretroviral agents on mtDNA synthesis in vivo and to assess the value of monitoring mtDNA in PBMCs to predict adverse events amongst these patients. MtDNA levels in PBMCs were significantly decreased in treatment-naive HIV-1-infected patients compared with healthy people. MtDNA levels were not only significantly correlated with CD4(+) T-cell count, but also inversely correlated with HIV-1 viral load. MtDNA levels in untreated patients and healthy controls were stable during the period of observation. On the other hand, amongst patients treated with regimens containing AZT/3TC or d4T/3TC, mtDNA increased during treatment and recovered to levels comparable to healthy controls. In contrast, mtDNA decreased immediately after the initiation of an AZT/ddC-containing regimen. We did not find a correlation between mtDNA levels and changes in clinical parameters. There was no significant difference in mtDNA levels between patients with and those without lipoatrophy. Furthermore, there was no obvious difference in mtDNA levels amongst those patients exhibiting signs and symptoms of peripheral neuropathy. In conclusion, the decrease in mtDNA levels in PBMCs amongst HIV-1-infected patients and its amelioration by antiretroviral therapy may suggest the influence of direct effects on mitochondria or mtDNA by HIV-1 infection. Further investigations are needed to elucidate the mechanisms contributing to decreased mtDNA and the value of mtDNA measurement in the care of HIV-1-infected individuals.     

A novel bioactive 31-amino acid ET-1 peptide stimulates eosinophil recruitment and increases the levels of eotaxin and IL-5

OBJECTIVE AND DESIGN: Investigation of the role of a novel inflammatory mediator 31-amino acid endothelin-1 [ET-1 (1-31)], a major ET derivative in granulocytes, in eosinophil recruitment after its subcutaneous administration to mice. METHODS: Various ET-1 derivatives (100 pmol), with or without ET receptor antagonists (200 pmol), were administered subcutaneously to mice, and then the eosinophil migration into and chemokine levels in the injected loci were analyzed. RESULTS: ET-1 (1-31) and a 21-amino acid endothelin-1 (ET-1), but not big ET-1, induced eosinophil migration into the injected loci with a peak after administration for 12 h, and increased the levels of eotaxin and interleukin-5 with peaks at 6 and 24 h, respectively. These effects of ET-1(1-31) and ET-1 were significantly inhibited by an ETA receptor antagonist, BQ-123, but not by an ETB receptor antagonist, BQ-788. CONCLUSION: Novel bioactive ET-1 (1-31) induces local eosinophil migration, and increases in eotaxin and interleukin-5 through an ETA or ETA-like receptor.     

Effects of vitamin A on proliferation of human distal airway epithelial cells in culture

Using a serum-free culture method, we investigated the effects of vitamin A on the proliferation of human distal airway epithelial cells. Outgrowth of epithelial cells from lung tissue explants was enhanced by treatment with all-trans retinol at concentrations of 10^–8 to 10^–7 M. The colony-forming activity of cells harvested from the primary culture and replated onto Swiss 3T3 fibroblastic feeders was, in contrast, significantly reduced by 10^–7 M to 10^–5 M retinol. When the primary cells were harvested and subcultured on Primaria plates, population expansion was also inhibited by retinol at 10^–10 to 10^–6 M. We further investigated the cells to determine whether there was any difference in sensitivity to the growth-inhibitory effects of vitamin A between cells from the primary culture incubated with and without retinol. The population increase in cells harvested from the primary culture was inhibited equally in retinol-treated and non-treated cells by subsequent treatment with retinol or retinoic acid, this inhibition being dose-dependent. DNA synthetic activity was also inhibited. Interestingly, both the growth rate and the colony-forming efficiency on feeders were greater in the subculture of cells from the retinol-treated primary culture than in those non-treated. When the cells in the secondary subculture were treated with retinoic acid and replated again, they showed a greater population increase rate than those non-treated. Our results showed that human distal airway epithelial cells isolated from lung tissue were sensitive to the growth-inhibitory effect of vitamin A, but the proliferative potential in some fraction of the epithelial cell population was possibly enhanced by vitamin A treatment.     

Persistence of archetypal JC virus DNA in normal renal tissue derived from tumor-bearing patients.

JC virus DNAs derived from the urine of nonimmunosuppressed individuals generally contain an archetypal regulatory region which may have generated various regulatory regions of JC virus from from the brain with progressive multifocal leukoencephalopathy (PML). In this study, we examined whether JC virus persisting in normal human kidney tissue contains the archetypal regulatory region. Renal medulla, cortex, and tumor from 32 patients bearing renal tumors were screened for JC virus DNA by blot hybridization. Viral DNA was detected in the medulla in 13 cases (41%), in the cortex in 2 cases (6%), but not at all from the tumor. A number of viral DNA-positive specimens (8 from the medulla and 2 from the cortex) were used to amplify and sequence viral regulatory regions by polymerase chain reaction. Structures of the regulatory regions from all the specimens were, with a few nucleotide variations, identical with that of the archetypal region which was previously detected in the JC virus DNA from urine. This finding supports the hypothesis that the JC virus associated with PML evolved from the archetypal JC virus during persistence in human hosts. Furthermore, we present evidence that renal JCV is replicating and that progeny virions are excreted into the urine.     

Expression of gastrin-releasing peptide (GRP) in the bovine uterus during the estrous cycle

Gastrin-releasing peptide (GRP) has been proposed as a novel regulatory peptide in the reproductive tract. We previously demonstrated that GRP immunoreactivities are found predominantly in the uterine gland epithelial cells of nonpregnant and pregnant cows. The present study focused on the distribution of GRP immunoreactivity and the expression of GRP mRNA in the bovine endometrium during the estrous cycle. Tissues were collected from 21 uterine horns and bodies during the estrous cycle. RT-PCR showed the expected GRP mRNA fragments (284 bp) in the tissues from all stages of the cycle. In situ hybridization results ascertained the expression of the GRP mRNA in the uterine gland epithelial cells and superficial epithelial cells of the endometrium. Positive staining of GRP immunoreactivity in the uterine gland epithelial cells was detected in both the uterine horn and body from all stages of the cycle. In metestrus and diestrus stages, GRP was also detected in the superficial epithelial cells of horn, but not in the body. The degrees of GRP mRNA expression and intensities of GRP immunoreactivity in the endometrium increased from proestrus to diestrus stages. These findings suggest that GRP may be important both in the endometrial remodeling during the estrous cycle and in the implantation and development of blastocysts.     

Cell to cell contact through ICAM-1-LFA-1 and TNF-alpha synergistically contributes to GM-CSF and subsequent cytokine synthesis in DBA/2 mice induced by 1,3-beta-D-Glucan SCG.

SCG is a major 6-branched 1,3-beta-D-glucan in Sparassis crispa Fr. showing antitumor activity. We recently found that the splenocytes from naive DBA/1 and DBA/2 mice are potently induced by SCG to produce interferon- gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), granulocyte-macrophage colony-stimulating factor (GM-CSF), and interleukin-12p70 (IL-12p70), and that GM-CSF plays a key biologic role among these cytokines. In this study, we investigated the contribution of cell-cell contact and soluble factors to cytokine induction by SCG in DBA/2 mice. Cell-cell contact involving intercellular adhesion molecule-1 (ICAM-1) and lymphocyte function-associated antigen-1 (LFA-1) was an essential step for the induction of GM-CSF and IFN-gamma by SCG but not for the induction of TNF-alpha or IL-12p70 by SCG. SCG directly induced adherent splenocytes to produce TNF-alpha and IL-12p70. GM-CSF was required for the induction of TNF-alpha by SCG, and in turn, TNF-alpha enhanced the release of GM-CSF and thereby augmented the induction of IL-12p70 and IFN-gamma by SCG. Neutralization of IL-12 significantly inhibited the induction of IFN-gamma by SCG. We concluded that induction of GM-CSF production by SCG was mediated through ICAM-1 and LFA-1 interaction, GM-CSF subsequently contributed to further cytokine induction by SCG, and reciprocal actions of the cytokines were essential for enhancement of the overall response to SCG in DBA/2 mice.     

Antigenicity of cryopreserved arterial allografts: comparison with fresh and glutaraldehyde treated grafts

The use of cryopreserved aortic allografts in cardiovascular surgery is widespread and has resulted in excellent outcomes. However, it is controversial whether cryopreservation suppresses the antigenicity of tissue. We designed experimental models to study whether the cryopreservation process alters antigenicity in comparison with that found in fresh and glutaraldehyde treated tissues. Fresh, cryopreserved, and glutaraldehyde treated thoracic aorta from Brown Norway rats were subcutaneously implanted into Lewis rats. Inflammatory cells infiltrating around the grafts were measured on days 7, 14, 28, and 56 after implantation. The glutaraldehyde treated grafts showed significantly less infiltration than the fresh or cryopreserved grafts (p < 0.005). No significant difference was detected between the fresh and cryopreserved grafts. Another study examined the effect of modifications of the aortic allograft on subsequent allogeneic skin graft antigenicity. Subcutaneous implantation of fresh, cryopreserved, and glutaraldehyde treated aortic grafts from Brown Norway into Lewis rats resulted in subsequent skin graft rejection at 4.4+/-0.7, 5.1+/-0.8, and 6.6+/-2.1 days, respectively. There was no significant difference between the fresh and cryopreserved groups; whereas skin grafts in the glutaraldehyde group survived longer than those in the cryopreserved group. These results indicate that cryopreservation had no significant influence on antigenic suppression of arterial allografts.     

Psychosocial factors regulating natural-killer cell activity in recurrent spontaneous abortions

PROBLEM: The preconceptional natural-killer cell (NK) activity predicts subsequent miscarriage among women with unexplained recurrent spontaneous abortion (RSA). Psycho-neuro-immuno-endocrine network has recently been proposed as a mechanism for abortions. We therefore examined which psychosocial factors influenced the NK activity among women with RSA. METHOD OF STUDY: We measured the preconceptional NK activity of 61 women with a history two consecutive unexplained first-trimester miscarriages and no live births. We also administered semi-structured interviews and a battery of self-report questionnaires to assess their social support, personality, self-esteem and psychiatric symptoms. RESULTS: The preconceptional NK activity was negatively correlated with the women's neuroticism personality trait (r= -0.32, P = 0.01) and current depressive symptoms (r = -0.26, P= 0.05), and positively correlated with their self-esteem (r = 0.34, P = 0.01). CONCLUSIONS: In addition to several substances such as transforming-growth-factor beta and granulocyte-macrophase colony-stimulating factor, we found that low neuroticism, low depression scale score and high self-esteem contributed to high NK activity among women with RSA.