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Background: We have recently shown that the expression of nestin, a progenitor/stem cell marker protein, is localized in different mesenchymal compartments in human skin including the sweat gland stroma., Objectives: As other exocrine glands are recognized sources of multipotent stem cell populations with potential for multilineage differentiation, it was our aim to isolate, expand and characterize glandular stem cells from human sweat glands., Methods: Isolation of human sweat glands was based on mechanical and enzymatic digestion of axillary skin. Cultivation was performed on collagen-coated cell culture dishes and the resulting cell population was investigated at the protein and mRNA level., Results: Outgrowing cells of isolated sweat glands showed a high-proliferation activity and were characterized by nestin expression in more than 80% of the cells. These sweat gland stem cells could be maintained in culture for long periods of time and showed spontaneous differentiation into cells representative of the different germ layers., Conclusions: This pilot study provides the first, simple protocol for the isolation of adult human nestin-positive stem cells from the sweat gland mesenchyme, which promises to provide an easily accessible and abundantly available, autologous source of multipotent stem cells for cell-based regenerative medicine applications.

Cerebral magnetic resonance imaging findings are of great value for the diagnosis of nonacquired GH deficiency (GHD), and ectopic posterior pituitary hyperintense signal (EPPHS) is a sensitive and specific indicator of hypopituitarism. It has been suggested that patients with childhood-onset GHD and EPPHS do not require additional investigation of GH secretion and should not be retested when adult height is achieved. This recommendation has never been validated through a systematic study. This study aimed to characterize the anterior pituitary function status of patients with EPPHS treated for GHD during childhood after completion of GH therapy when adult height had been achieved. Patients (n = 18; 15 males and three females) with childhood-onset GHD associated with ectopic neurohypophysis were treated with hGH (0.20 +/- 0.05 mg/kg.wk) for 9.9 +/- 4.0 yr (from 6.8 +/- 4.7 to 17.7 +/- 1.3 yr of age) with a mean height gain of 2.6 +/- 1.4 sd score. GH secretion was reevaluated by arginine insulin (n = 15) or propanolol glucagon (n = 3) test after 0.5 +/- 0.6 yr of GH withdrawal. At reevaluation, peak GH was more than 10 mug/liter in four patients (22%; range, 11.7-19.5 microg/liter; group I), between 5 and 10 microg/liter in three patients (17%; range, 7.3-9 mug/liter; group II), and less than 5 microg/liter in 11 patients (61%; range, 0-4.7 microg/liter; group III). A positive correlation was found between serum IGF-I and peak GH levels after attainment of adult height (P = 0.007). Only one of the seven patients who showed increased GH secretion ability in adulthood (groups I and II) demonstrated other hormonal deficiencies (gonadotropin and adrenal insufficiencies). Among the 11 patients with persistent severe GHD (group III), 10 (91%) of the 11 subjects were shown to have multiple pituitary hormone deficits after attainment of adult height. The structure of the hypothalamo-pituitary axis differs among groups [i.e. patients who showed increased GH secretion ability in adulthood (groups I and II) vs. those who remained severely GHD (group III)]. The location of the EPPHS was significantly different among groups (P < 0.003). The EPPHS was found at the median eminence in all but one of group III patients and along the pituitary stalk (proximal stalk) in all but one of group I and II patients. The pituitary stalk was visible and described as normal (n = 1) or thin (n = 6) in all group I and II patients, whereas the pituitary stalk was not visible even after enhancement in seven of the 11 group III patients (P < 0.02). The prevalence of anterior pituitary hypoplasia and the mean height gain sd score were similar in each group. In conclusion, only 61% of patients with childhood-onset GHD and EPPHS remained severely GHD, and thus suitable for GH therapy, in adulthood. Although the pathogenesis of anterior pituitary dysfunction remains unclear in patients with ectopic neurohypophysis, isolated GHD, location of EPPHS along the stalk, and visibility of the pituitary stalk on magnetic resonance imaging findings clearly represent important markers to predict a less severe form of the disease.

Two human immunodeficiency virus-infected patients with refractory disseminated Mycobacterium avium complex infection were treated with recombinant interferon-gamma (IFN-gamma) given subcutaneously for 3 and 4 months, respectively. Although both patients demonstrated some clinical improvement initially, IFN-gamma therapy did not produce sustained benefit.

Liver enzyme elevation (LEE) is commonly observed after combination antiretroviral therapy (ARVT) for HIV infection is begun. Potential risk factors for LEE after treatment with ritonavir and saquinavir with or without stavudine were investigated in 208 HIV-infected patients, by use of the Cox proportional hazard model. Eighteen patients (9%) developed LEE during the 48-week follow-up. Multivariate analysis, adjusted for baseline levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), showed that hepatitis B surface antigen (HBsAg) positivity (relative risk [RR], 8.8; 95% confidence interval [CI], 3.3-23.1) and the use of stavudine (RR, 4.9; 95% CI, 1.5-16.0) were the only significant risk factors for developing LEE. After LEE occurred, ALT and AST concentrations decreased by >50% in 13 of 14 patients who continued ARVT during LEE. In this study, it appeared safe to continue ARVT during LEE; however, more data from larger studies are required to confirm this finding.

Today's antiretroviral combination regimens can induce significant and sustained decreases in human immunodeficiency virus (HIV)-RNA levels, allowing the immune system to recover. To what extent immune reconstitution is possible and what factors determine the outcome have thus far not been resolved. We studied 19 subjects, treated for 2 years with protease inhibitor-containing triple therapy, who had a strong suppression of HIV-RNA levels. CD4+ T-cell numbers increased from medians of 170 to 420x106 cells/L, but in a number of subjects T-cell numbers did not further increase after week 72, without having reached normal values. Long-term CD4+ T-cell change was mainly caused by a slow but continuous increase in naive CD4+ T cells (CD45RA+CD62L+) and was predicted by the baseline number of these cells. Our data indicate that long-term immunological recovery is gradual, even during strong suppression of viral replication, not always complete, and dependent on the preexisting level of naive CD4+ T cells.

Lymphoproliferative responses (LPRs) to recall antigens (Ags) and human immunodeficiency virus type 1 (HIV-1) Gag and frequencies of circulating HIV-1-specific cytotoxic T lymphocyte precursors (CTLps) were measured in 12 patients undergoing highly active antiretroviral therapy (HAART) after long-standing HIV-1 infection. LPRs to at least 1 recall Ag became detectable or increased in all patients during HAART. No significant LPRs to Gag-p24 were observed, whereas 4 of 8 patients tested presented with Gag-p17-specific LPRs. HIV-1-specific CTLp frequencies became measurable or increased early during therapy in 6 of 10 patients tested and were maintained or decreased thereafter. Increasing HIV-1-specific CTLp frequencies were seen only in association with partial HAART failure in 1 patient. In conclusion, restoration of CD4+ T lymphocyte responsiveness to recall Ags is achieved during HAART. The data provide evidence for limited HIV-1-specific CD4+ memory T cells during advanced HIV-1 infection and suggest that both CD4+ and CD8+ HIV-1-specific T cells are poorly stimulated when viral load is suppressed.

Vascular endothelial growth factor (VEGF) is an endothelial cell specific mitogen that has been implicated in hypoxia-mediated angiogenesis under physiological and pathological conditions. We used the middle cerebral artery occlusion model (MCAO) in the rat to investigate VEGF mRNA and protein localization, and VEGFR-1 mRNA and VEGFR-2 mRNA expression in cerebral ischemia. By nonradioactive in situ hybridization we observed upregulation of VEGF mRNA and VEGFR-1 mRNA, but not of VEGFR-2 mRNA in the hemisphere ipsilateral to MCA occlusion. VEGF mRNA was upregulated in the periphery of the ischemic area commencing 3 hours (h) after onset of MCAO, reached a peak after 24 h, and remained expressed at lower levels until 7 days (d) after MCAO. Double labelling experiments revealed that the majority of VEGF expressing cells in the penumbra and within the infarct were immunoreactive for Ox-42, Iba-1, and Ed1, but not for GFAP and neurofilament proteins, suggesting that microglial cells/macrophages are the major cell type expressing VEGE Since VEGF was also expressed in Ox-42 immunoreactive cells distant from the infarct (e.g. in the corpus callosum and hippocampus), activated microglial cells expressing VEGF may migrate towards the ischemic stimulus. VEGF protein was also detected on capillaries within the peri-ischemic area, suggesting that VEGF produced and secreted by microglial cells/macrophages binds to its receptors on nearby vascular endothelial cells and initiates an angiogenic response which counterbalances tissue hypoxia. Accordingly, apoptosis of neuroectodermal cells in the penumbra was highly depressed after the onset of angiogenesis. The spatial and temporal correlation between the induction of angiogenesis with VEGF and VEGFR-1 expression suggests that the ischemic upregulation of VEGF represents a physiological response of the brain to counterbalance hypoxia/ischemia in order to protect neuroectodermal tissue.

Background: A study was conducted to estimate the incidence of health problems in HIV-infected travelers with various degrees of immunodeficiency to the (sub)tropics., Methods: A retrospective questionnaire-based study among HIV-infected patients attending the outpatient department of a university hospital during three months in 1996 with a history of travel to (sub)tropical destinations in the proceeding 12 months. The outcome measures were incidences of and medical consultation rates for common travel-associated illnesses., Results: Of 293 HIV-infected patients, 59 (20%) had traveled during the preceding 12 months; of these 36 (61%) responded. Ten (28%) had traveled more than once during this period. There were no significant differences between travelers and nontravelers regarding CD4 count and age. Fifteen respondents (42%) used cotrimoxazole (CTX) for PCP prophylaxis; 22 (61%) had sought pretravel health advice. Median duration of travel was 3 weeks. Respiratory infections were recorded by two respondents (6%, 95% confidence intervals 1-19%) and skin problems by 10 (28%, 14-45%). Of 31 respondents without diarrhea on departure, 10 recorded diarrhea (32%, 19-57%). The overall rate of medical consultation for travel-related complaints ws 5% abroad and 28% after return, respectively. There was no association between the risk of diarrhea and CD4 count or CTX prophylaxis, even after adjustment for differences in age, duration of travel and travel experience., Conclusions: Although numbers in this study are small, HIV-infected travelers tend to have a high rate of medical consultation for (possibly) travel-related illness. Diarrhea is the most frequent complaint, but shows no strong association with degree of immunodeficiency.

The outcome and predictors of virologic treatment failure of highly active antiretroviral therapy (HAART) were determined for 271 human immunodeficiency virus (HIV)-infected protease inhibitor-naive persons. During a follow-up of 48 weeks after the initiation of HAART, 6.3% of patients experienced at least one new AIDS-defining event, and 3.0% died. Virologic treatment failure occurred in 40% (indinavir, 27%; ritonavir, 30%; saquinavir, 59%; ritonavir plus saquinavir, 32%; chi2, P=.001). Risk factors for treatment failure were baseline plasma HIV-1 RNA (odds ratio [OR], 1.70 per log10 copies increase in plasma HIV-1 RNA), baseline CD4 cell count (OR, 1. 35 per 100 CD4 cells/mm3 decrease), and use of saquinavir versus other protease inhibitors (OR, 3.21). During the first year of treatment, 53% of all patients changed (part of) their original HAART regimen at least once. This was significantly more frequent for regimens containing saquinavir (62%; 27% for virologic failure) or ritonavir (64%; 55% for intolerance) as single protease inhibitor.

The presence of syncytium-inducing (SI) human immunodeficiency virus type 1 (HIV-1) variants is predictive for accelerated progression to AIDS. This study showed that a 4-year survival with AIDS also occurred significantly more often for patients who lacked SI variants. However, multivariate Cox analysis excluded the predictive value of SI viruses for rapid death as being independent from low CD4+ T cell counts. Incidence of appearance of SI variants was increased in persons with CD4+ T cell counts <500/microliter but remained constant in the strata of CD4+ T cell counts <500/microliter, excluding the possibility that loss of immune control is the only prerequisite for the development of SI HIV-1 variants.

Anti-human immunodeficiency virus type 1 (HIV-1) activity was assessed in HIV-1-infected homosexual and bisexual men receiving 18-36 MIU/day of recombinant interferon (IFN)-alpha2a for Kaposi's sarcoma (KS). The median baseline HIV-1 RNA level was 4.99 log10 copies/mL. Seventeen subjects (68%) showed an RNA decline > or = .5 log10/mL, with a maximum at week 4 (median decline = 1.91, range = 3.64-1.15; P = .0007), after which RNA levels stabilized. Eight subjects (32%) with lower median initial CD4+ T cell counts (60 vs. 350 x 10(6)/L; P = .01) did not show RNA responses. Neither RNA nor KS responses were negatively affected by IFN-alpha2a dose modifications. Anti-HIV responses of KS responders (n = 15) and nonresponders (n = 10) did not differ. High-dose IFN-alpha can exert potent anti-HIV activity that is not associated with anti-KS activity.

Exposure of human immunodeficiency virus to the nucleoside analogue lamivudine (3TC) rapidly selects for resistant variants with a valine at codon 184 (M184V) in the catalytic site of reverse transcriptase. In vitro, 184V demonstrated increased enzyme fidelity and suppressed zidovudine resistance. Clinical trials demonstrated that 3TC-zidovudine combination therapy results in a strong and sustained antiviral response. To investigate the role of 184V on in vivo virus evolution, the effect of zidovudine addition in 3TC-pretreated patients harboring 184V was studied. In vivo, no significant change in fidelity was observed with 184V, shown by generation of the classical pattern of zidovudine mutations. Of interest, in contrast to zidovudine monotherapy, in which just one substitution is sufficient for in vivo development of significant zidovudine resistance, multiple substitutions are required for the same level of zidovudine resistance in strains harboring 184V. This need for multiple substitutions may be one of the mechanisms explaining the sustained antiretroviral response of the 3TC-zidovudine combination.

We performed a cross-sectional study at an outpatient AIDS clinic to assess the prevalence of Campylobacter species in stool specimens from 201 consecutive patients infected with human immunodeficiency virus (HIV). We characterized campylobacters phenotypically and genetically by using primers for the group of common species (i.e., C. jejuni, C. coli, C. lari, and C. upsaliensis) and for most individual uncommon species. We performed cultures with use of a membrane filter technique on nonselective blood agar and found that Campylobacter species were the most frequent enteropathogenic bacteria: the organisms were recovered from 7 (16%) of 43 patients with diarrhea and 5 (3%) of 158 patients without diarrhea (P = .001). We isolated only one campylobacter with use of conventional culture techniques on selective media. Phenotypic characterization of 10 campylobacter strains resulted in the misidentification of four isolates. C. upsaliensis was the most frequently isolated species, followed by C. jejuni and C. coli. Two strains could not be identified with the available primers. Two of 12 Campylobacter strains were resistant to erythromycin, and two were resistant to ciprofloxacin. We conclude that Campylobacter species other than C. jejuni can frequently be detected in the stools of HIV-infected patients and that these organisms could be associated with diarrhea.

To study the effect of sequential or additive use of zalcitabine or didanosine on survival in 308 human immunodeficiency virus-infected patients with advanced disease treated with zidovudine, an observational study using time-dependent Cox proportional hazards models was done. Changing to sequential or additive therapy was based on deterioration of a patient's health status, a significant drop in CD4 cell count, or intolerance for zidovudine. The median CD4 cell count at baseline was 110 x 10(6)/L; 42% of patients had AIDS. The median count before a change in therapy was 50 x 10(6)/L. Additive or sequential treatment was associated with an increased risk for death (relative hazard, 1.59; 95% confidence interval [CI], 1.01-2.49; and 1.58; 95% CI, 1.10-2.37, respectively). Adjustment of the models for prognostic factors failed to substantially affect this observation. Possibly the lack of benefit in this study is because patients switched therapy at advanced stages, whereas the switch may be more effective in early disease.

This study examined the factors associated with the development of a first episode of Pneumocystis carinii pneumonia (PCP) in 5,025 patients with AIDS, including 1,976 patients with primary PCP at the time of AIDS diagnosis and 635 with primary PCP occurring subsequently. Compared with untreated patients, patients treated with zidovudine were at similar risk of developing PCP during the first year of therapy but were at greater risk after longer intervals of treatment. The following factors were associated with an increased risk of PCP (either at the time of AIDS diagnosis or thereafter): lack of primary PCP prophylaxis, male homosexuality/bisexuality, diagnosis of AIDS in northern Europe, and CD4 cell count below 200 x 10(6)/L at the time of AIDS diagnosis. Patients with severe weight loss had a 60% higher risk of developing PCP during follow-up than those without such weight loss. Thus, the occurrence of PCP depended on geographic location, mode of acquisition of human immunodeficiency virus and AIDS, degree of immunodeficiency, and use of various treatment regimens.

The effect of the appearance of drug-resistant human immunodeficiency virus type 1 (HIV-1) on viral RNA load was studied in patients treated with the reverse transcriptase inhibitor lamivudine. During the first 12 weeks of treatment, HIV-1 RNA concentrations and amino acid changes in codon 184, causing high-level resistance to lamivudine, were determined in longitudinal serum samples from HIV-1 p24 antigen-positive and -negative patients. A marked decline in the amount of HIV-1 RNA (approximately 95% below baseline) and HIV-1 p24 antigen was observed within 2 weeks, followed by a rise that coincided with the appearance of lamivudine-resistant viruses in serum (isoleucine mutants initially, which were subsequently replaced by valine variants). After 12 weeks, a partial antiviral effect was observed despite the presence of a complete codon 184 mutant virus population in serum. This study shows that the rapid appearance of drug-resistant virus in serum is followed by an increase in viral RNA load.

We investigated the in vitro effect of cultured human thymic epithelial monolayer cells on mononuclear cells (MNC) from patients with Acquired Immune Deficiency Syndrome (AIDS) and AIDS-related complex (ARC). Patients having undergone bone-marrow transplantation (BMT), who showed a similar deficiency of T-lymphocytes expressing CD4 (Leu-3, T4), and healthy blood donors served as controls. Most epithelial monolayer cells were of thymic medulla origin, as documented using a panel of monoclonal antibodies to thymic epithelial cells. In AIDS/ARC patients the CD4-positive cells (ranging between 4 and 30% of MNC) increased by a factor of 1.56 (s.e.m. 0.15, n = 12) during a 2 h incubation on the monolayer. Human fetal lung fibroblasts were inactive in this respect. There was no consistent change in cells expressing CD3 (Leu-4, T3), CD5 (Leu-1) or CD8 (Leu-2, T8). In BMT patients and healthy controls, neither thymus epithelium nor fibroblasts had any effect on T cell marker expression. Incorporation of tritiated thymidine by stimulated or unstimulated lymphocytes was increased after incubation for 3 days on either epithelium or fibroblasts. We conclude that CD4 expression is induced on MNC from AIDS/ARC patients during a 2 h incubation on epithelial monolayers.