Human immunodeficiency virus infection of eosinophils in human bone marrow cultures.

Normal human bone marrow, cultured in vitro with interleukin 5 to promote eosinophil production and maturation, was inoculated with cell-free isolates of human immunodeficiency virus type 1 (HIV-1). CD4 expression by eosinophil precursors, determined by immunocytochemistry, was found to be greatest early in their maturation with a rapid decline after 28 d in culture. Productive HIV infection of eosinophil precursors was detected 14 d after inoculation, by a combination of immunostaining for HIV-1 p24 and gp41/160 and in situ hybridization for viral RNA, together with assay of culture supernatants for p24 antigen and reverse transcriptase activity. Thus, eosinophils are susceptible to productive HIV-1 infection in vitro and may be an important reservoir for the virus in vivo.

Detection of plasma tumor necrosis factor, interleukins 6, and 8 during the Jarisch-Herxheimer Reaction of relapsing fever.

The Jarisch-Herxheimer Reaction (J-HR) is a clinical syndrome occurring soon after the first adequate dose of an antimicrobial drug to treat infectious diseases such as Lyme disease, syphilis, and relapsing fever. Previous attempts to identify factors mediating this reaction, that may cause death, have been unsuccessful. We conducted a prospective trial in Addis Ababa, Ethiopia on 17 patients treated with penicillin for proven louse-borne relapsing fever due to Borrelia recurrentis to evaluate the association of symptoms with plasma levels of tumor necrosis factor (TNF), interleukins 6, and 8 (IL-6 and -8). 14 of the 17 (82%) patients experienced a typical J-HR consisting of rigors, a rise in body temperature (1.06 +/- 0.2 degrees C) peaking at 2 h, leukopenia (7.4 +/- 0.6 x 10(-3) cells/mm3) at 4 h, a slight decrease, and then rise of mean arterial blood pressure. Spirochetes were cleared from blood in 5 +/- 1 h after penicillin. There were no fatalities, but constitutional symptoms were severe during J-HR. Plasma TNF, IL-6, and -8 were raised in several patients on admission, but a seven-, six-, and fourfold elevation of these plasma cytokine concentrations over admission levels was detected, respectively, occurring in transient form coincidental with observed pathophysiological changes of J-HR. Elevated plasma cytokine levels were not detected in the three patients who did not suffer J-HR. We conclude that the severe pathophysiological changes characterizing the J-HR occurring on penicillin treatment of louse-borne relapsing fever are closely associated with transient elevation of plasma TNF, IL-6, and -8 concentrations.

Acute systemic inflammation impairs endothelium-dependent dilatation in humans.

BACKGROUND: We tested the hypothesis that endothelial dysfunction underlies the association between an acute inflammatory episode and the transiently increased risk of a cardiovascular event by examining the effects of an experimental inflammatory stimulus on endothelium-dependent vasodilation. METHODS AND RESULTS: Salmonella typhi vaccine was used to generate a systemic inflammatory response in healthy volunteers. In 12 subjects, dilatation of the brachial artery to flow and to sublingual nitroglycerin (NTG) was recorded (conduit vessel response), and in 6 subjects, venous occlusion plethysmography was used to measure forearm blood flow during intrabrachial infusion of the endothelium-dependent dilators acetylcholine (ACh) and bradykinin (BK) and the endothelium-independent dilators NTG and verapamil (resistance vessel response). Responses were assessed 16 hours before and 8 and 32 hours after vaccination. Vaccination resulted in elevations in white cell count and serum levels of interleukin-6 and interleukin-1 receptor antagonist. Eight hours after vaccination, resistance vessel responses to BK (P:=0.0099) and ACh (P:=0.0414) were markedly attenuated, and brachial artery flow-mediated dilatation was depressed. Resistance vessel responses to verapamil and NTG were unchanged, as was the conduit vessel response to NTG. Thirty-two hours after vaccination, resistance vessel responses to BK and ACh had returned to normal. CONCLUSIONS: S typhi vaccine generates a mild inflammatory reaction associated with temporary but profound dysfunction of the arterial endothelium in both resistance and conduit vessels to both physical and pharmacological dilator stimuli. This finding might explain the association between infection and inflammation and the enhanced risk of an acute cardiovascular event.

Autonomic denervation in jejunal mucosa of homosexual men infected with HIV.

Autonomic nerves in jejunal mucosa of HIV-infected patients show severe structural damage on electron microscopic examination. The aim of this study was to quantify loss of autonomic axons from the lamina propria of HIV-infected patients in different clinical stages of disease. Jejunal biopsies were taken from 19 HIV-antibody-positive homosexual men and from 10 control patients. Autonomic fibres in the mucosa were stained with a neurone-specific polyclonal antibody, PGP 9.5. The density of axons was quantified by a point-counting technique using a Lennox eyepiece graticule under light microscopic examination. There was significant reduction in axonal density in the villi of HIV-infected patients [mean, 9.0; standard deviation (s.d.), 4.7] compared with controls (mean, 15.3; s.d., 5.2; P = 0.003), and in the pericryptal lamina propria of HIV-infected patients (mean, 17.8; s.d., 5.4) compared with controls (mean, 27.3; s.d., 6.2; P = 0.0002). Although autonomic denervation occurs throughout the jejunal mucosa of HIV-infected patients, there was no correlation between the clinical stage of HIV disease and the degree of denervation. The denervation was greatest in patients with the most severe diarrhoea, but this difference was not significant. This study provides the first quantitative morphological evidence for depletion of autonomic nerves in the jejunum of patients infected with HIV. Autonomic neuropathy may contribute to chronic diarrhoea in HIV disease.

Direct in vitro infection of human intestine with HIV-1.

OBJECTIVE: To directly infect human fetal intestine with HIV in vitro. DESIGN: Human fetal intestinal explant cultures were exposed to HIV-1 and monitored for evidence of infection by biochemical assay, immunohistochemistry and in situ hybridization. METHODS: Human fetal intestinal explants (14-21 weeks) were established in culture and exposed to HIV-1. Tissue culture fluid was assayed for p24 antigen and reverse transcriptase activity over a 14-day period. Explants were removed from culture on days 4, 7, 10 and 14 postinoculation and subjected (1) to immunohistochemistry to detect p24 and gp160/41 antigens, and (2) to in situ hybridization to detect HIV-1 RNA. Explant tissue culture fluid was cocultured with Jurkat T-cells to detect infectious viral particles. RESULTS: Reverse transcriptase activity and p24 antigen levels in fetal explant culture fluid rose between 7 and 14 days after viral inoculation. Jurkat T-cell cocultures confirmed the presence of infectious virus. Cells in the lamina propria resembling lymphocytes and macrophages of both small intestine and colon stained positively for the viral proteins p24 and gp41. The same type of cells also stained positively for HIV-1 RNA using in situ hybridization. Dual-label immunohistochemistry, combined immunohistochemistry and in situ hybridization confirmed the presence of viral protein and RNA in cells bearing the CD3, CD4 (lymphocyte) or CD68 (macrophage) surface markers. There was no evidence at any time of HIV-1 infection of epithelial cells. CONCLUSIONS: Cells of the lamina propria of the small intestine and colon, bearing lymphocyte or macrophage markers, can be directly infected by and support the replication of HIV-1. Such infection may be implicated in the pathogenesis of HIV enteropathy.

HIV infection of human fetal intestinal explant cultures induces epithelial cell proliferation.

OBJECTIVE: The concept that HIV infection per se alters small intestinal mucosal structure and function (HIV enteropathy) remains controversial and in this study we report in vitro experiments designed to elucidate the matter. METHODS: Twenty pairs of human fetal intestinal tissue explants were maintained in vitro for up to 14 days; one explant of each pair was incubated and infected with HIV, and the other served as a matched uninfected control. At various times after infection, explant culture fluid and tissue were removed, p24 concentration was measured and tissue formalin fixed. Explant tissue was embedded in paraffin wax and sections stained by an immunoperoxidase method directed against proliferating cell nuclear antigen (PCNA). The percentage of proliferating crypt and villous epithelial cells, stained by PCNA, was calculated in paired samples. The difference between the percentage for paired samples was designated delta crypt proliferation (delta CP) and delta villous proliferation (delta VP), respectively. Epithelial cell proliferation was deemed to be enhanced if the percentage of PCNA-stained cells was greater in the HIV-infected than in the control tissue. RESULTS: Explant culture fluid from tissue exposed to HIV showed a progressive rise in p24 antigen (Ag) level, indicating HIV infection of these explants. Fifteen pairs of explants showed progressively positive delta CP with time (P < 0.01) indicating crypt hyperplasia and all 20 pairs of explants showed positive delta VP, indicating hyperplasia of villous epithelial cells. CONCLUSIONS: This study provides direct evidence that HIV stimulates epithelial cell proliferation in intestinal mucosa. HIV-infected human intestinal explants provide a model of crypt hyperplastic villous atrophy previously described as HIV enteropathy and detected in clinical biopsy specimens from HIV-infected patients.

Immune response following oral administration of cholera toxin B subunit to HIV-1-infected UK and Kenyan subjects.

OBJECTIVE: To determine the effect of HIV-1 infection on immunoglobulin (Ig) G and IgA antibody response and circulating antibody forming cell response to oral immunization with the B subunit of cholera toxin. DESIGN: Healthy UK volunteers, and HIV-1-positive UK and Kenyan volunteers at different clinical stages of HIV-1 infection received two oral immunizations. CD4+ T cells, serum beta 2-microglobulin and neopterin were measured as surrogate markers of disease stage, and correlated with immunization response. METHODS: Serum antitoxin IgG and IgA measured by enzyme-linked immunosorbent assay and antitoxin IgG, IgA and IgM antibody-forming cells detected by enzyme-linked immunospot assay at different times after two oral immunizations. RESULTS: UK HIV-positive volunteers (mean CD4+ T cell count, 52 x 10(6)/l) responded poorly to primary and booster immunization. HIV-infected Kenyans (752 x 10(6)/l CD4+ T cells) had a significant primary and booster antibody response, whereas those with a mean CD4+ T cell count 186 x 10(6)/l had an insignificant primary, but significant booster response. Two oral immunizations induced antibody responses in HIV-positive Kenyan groups (who may have prior immunity from exposure to environmental bacterial toxins) of similar or greater magnitude to healthy UK volunteers. CONCLUSIONS: Mucosal immunization may recall immune memory and be of benefit in early and moderately advanced clinical HIV disease. The findings have important clinical implications in that mucosally targeted vaccines are potentially useful in this group of patients.

Damage to jejunal intrinsic autonomic nerves in HIV infection.

Jejunal biopsies from 11 HIV-infected male homosexuals, without secondary enteropathogen infection, were examined at ultrastructural and light microscope level. Subjects were clinically categorized into four groups: asymptomatic (3), AIDS-related complex (4), persistent generalized lymphadenopathy (1), and AIDS (3). All 11 biopsies, including the three from asymptomatic HIV-infected individuals, showed extensive damage to autonomic nerve fibres in the lamina propria. These findings show that HIV infection damages the enteric autonomic nerves. Since asymptomatic HIV-infected individuals had similar damage, this appears to be an early event in the course of HIV infection.

Impaired absorption of zidovudine in patients with AIDS-related small intestinal disease.

OBJECTIVES: To investigate the effect of small intestinal disease (SID) on the absorption of zidovudine (ZDV) in patients with AIDS. METHODS: Fourteen fasted homosexual men with AIDS received a single oral dose of ZDV (5 mg/kg). Nine subjects had clinical evidence of intestinal disease (chronic diarrhoea with wasting) confirmed by reduced fat absorption measured indirectly using the 14C-triolein test. Five subjects had AIDS-related symptoms other than those affecting the gastrointestinal tract with normal fat absorption. Sequential measurements of plasma ZDV including its glucuronide metabolite (GZDV) were obtained using radio-immunoassay and ZDV/GZDV concentrations-time profiles of both groups of subjects were compared. Comparisons were also made for each of the following computed variables: the maximum plasma concentration (Cmax), time to reach Cmax (Tmax), area under the plasma concentration-time curve (AUC0-6 h), the elimination half-life (t 1/2), and apparent oral clearance (CL0). RESULTS: In patients with SID, Cmax ZDV was reduced (6.39 +/- 3.39 versus 11.51 +/- 5.01 mumol/l; P < 0.05) and Tmax ZDV prolonged (0.81 +/- 0.51 versus 0.40 +/- 0.14 h; P < 0.05) but AUC0-6 h ZDV was no different from the non-SID group (8.03 +/- 2.73 versus 14.56 +/- 9.0 mumol/l-1xh; P = 0.06). There were no differences in t 1/2 ZDV (1.22 +/- 0.20 versus 1.13 +/- 0.30 h) or CL0 ZDV (3017 +/- 1158 versus 1700 +/- 889 ml/min; P > 0.05) between SID and non-SID groups, respectively, and GZDV values were comparable between the two groups. CONCLUSIONS: These data suggest delayed absorption rather than altered metabolism of ZDV in AIDS-related SID and raise the possibility of drug malabsorption. The clinical efficacy of orally administered low-dose ZDV regimens may require further evaluation in patients with chronic diarrhoea and AIDS.

Sustained plasma TNF-alpha and HIV-1 load despite resolution of other parameters of immune activation during treatment of tuberculosis in Africans.

OBJECTIVE: To determine the impact of treatment of tuberculosis on plasma HIV-1 load in African subjects and to correlate viral load with response to treatment and changes in immune activation. DESIGN: Clinical and microbiological responses, immune activation parameters and plasma HIV-1 load were determined in 20 patients with pulmonary tuberculosis and HIV-1 coinfection in Ghana, West Africa during the first 3 months of anti-tuberculosis treatment. METHODS: Plasma HIV-1 load and markers of immune activation were determined by commercially available assays. Human leukocyte antigen (HLA)-DR incorporation into the HIV-1 envelope was measured by using an immunomagnetic capture technique. RESULTS: Treatment of tuberculosis resulted in significant improvements in weight and haemoglobin, a high sputum smear conversion rate and marked reductions in mean plasma tumour necrosis factor (TNF) receptor-1, interleukin-6 and C-reactive protein. Furthermore, incorporation of host HLA-DR into the HIV-1 envelope decreased; this also suggested a reduction in immune activation of the cells supporting viral replication. However, of importance with regard to AIDS pathogenesis, neither mean plasma TNF-alpha nor HIV-1 load decreased significantly. CONCLUSIONS: The failure of HIV-1 plasma load to decline significantly during the initial months of anti-tuberculosis treatment is associated with high, sustained systemic levels of TNF-alpha. The dissociation between the sustained levels of plasma TNF-alpha and the major reductions in other, diverse immune activation parameters may represent dysregulation of cytokine production in these African patients.

Short-term growth hormone administration at the time of opportunistic infections in HIV-positive patients.

OBJECTIVES: A 12-week course of recombinant human growth hormone is an effective but expensive therapy for established HIV-related wasting. Wasting in HIV disease is often episodic, coinciding with bouts of acute opportunistic infection. We hypothesized that a short course of growth hormone, targeted at the time of opportunistic infection, might improve protein metabolism thereby reducing lean tissue loss. METHODS: HIV-infected men with acute opportunistic infections, who received standard antimicrobial treatment for their infection as well as intensive nutritional counselling and oral energy supplements, were randomized to receive growth hormone or placebo for 14 days. Principal assessments were protein metabolism (measured by 13C-leucine infusion), body composition (measured by DEXA) and safety. RESULTS: There were no significant changes in outcome parameters in the placebo group (n = 11). In the growth hormone group (n = 9), protein catabolic rate decreased by 60% in the fasted state (P = 0.02 versus placebo), lean body mass increased by 2.2 kg (P = 0.03 versus baseline) and fat mass decreased by 0.7 kg (P = 0.002 versus baseline). There was no increase in adverse or serious adverse events in the growth hormone as compared with the placebo group. CONCLUSIONS: A two-week course of growth hormone at the time of acute opportunistic infection in HIV-infected patients improves protein metabolism and body composition during therapy and appears to be safe. This may represent a rational and economical approach to the use of growth hormone therapy.

Increase in plasma viral load after oral cholera immunization of HIV-infected subjects.

OBJECTIVE: Constant antigenic stimulation of the large immune cell population contained within gut-associated lymphoid tissue during HIV infection may contribute to patients' total viral load. The aim of this investigation was to evaluate the effect of a mucosal antigenic challenge on HIV replication. DESIGN: Prospective clinical study. METHODS: Twelve HIV-1-infected men (mean age, 42.3 years) from the Casa de Apoio Santo Antonio, Rio de Janeiro, Brazil, were immunized with combined whole cell-toxin B subunit oral cholera vaccine. Blood was collected on days 0, 2, 4, 6, 10 and 15 after immunization and plasma was tested for cholera toxin-specific antibody response (IgG and IgA), beta2-microglobulin, and plasma viral load. CD4 lymphocyte counts were performed within 1 week before immunization. Five HIV-infected non-immunized individuals were studied as controls. RESULTS: There were no adverse effects following immunization and no deterioration in clinical outcome during 3 months of follow-up. A transient increase in viral load that ranged from twofold to 60-fold was observed in all cases and was statistically significant on days 2, 6 and 10 (P = 0.017, P = 0.025, P = 0.021, respectively). There was no correlation with CD4 cell counts. None of the non-immunized subjects demonstrated the pattern of viraemia observed after immunization (P > 0.10 on all days). CONCLUSIONS: Our data indicate that mucosal immunization with oral cholera vaccine induces a transient increase in HIV viraemia, regardless of clinical stage of infection and CD4 cell counts. These findings suggest that mucosal stimulation of HIV-infected patients enhances HIV replication.

The effect of endotoxin on skeletal muscle protein gene expression in the rat.

Sepsis is associated with net breakdown of skeletal muscle protein, mediated partly by reduced rates of muscle protein synthesis. This study investigated the role of altered gene expression for specific muscle proteins in mediating reduced protein synthesis in a rat model of acute severe sepsis. Adult rats were given a single sublethal intraperitoneal dose of endotoxin (bacterial lipopolysaccharide). Protein, RNA and DNA contents of muscle were measured and changes in expression of mRNA in tibialis anterior and extensor digitorum longus muscles were detected by quantification of Northern blots at 6, 24, 48 and 72 hr after endotoxin and in animals starved for 24 hr. Results showed that at 24 hr after endotoxin there was a loss of about 14% of muscle protein content. No reduction in mRNA was found at any time point for beta-myosin heavy chain (MHC), fast-MHC, alpha-actin, skeletal muscle troponin or carbonic anhydrase III (CA III); rather, at 48 hr there was increased expression of beta-MHC (224 +/- 123% control) and CA III (202 +/- 56%). Blocking TNF-alpha by pre-treatment with a monoclonal antibody did not appear to influence this. Total RNA content of muscle was reduced to 67% of the control values 24 hr after LPS, although this was no different to pair-fed animals starved for 24 hr. It is concluded that reduced protein synthesis in skeletal muscle in early acute sepsis is not primarily associated with reduced muscle protein gene expression.

Prospective analysis of patterns of weight change in stage IV human immunodeficiency virus infection.

Weight loss is a major manifestation of infection with the human immunodeficiency virus (HIV). Prospective analysis of weight change was performed in 30 male subjects with stage IV HIV infection over a period of 9-49 mo and weight change events (> 4 kg) related to contemporaneous clinical events. Two distinct patterns of weight loss were observed: episodes of acute severe weight loss and episodes of chronic unremitting progressive weight loss. Thirty-three acute episodes (median 9.1 kg in 1.7 mo) and 23 chronic episodes (13.2 kg in 9.5 mo) were identified. Twenty-seven of 33 (82%) acute weight-loss episodes were associated with nongastrointestinal opportunistic infections and 15 of 23 (65%) chronic episodes with gastrointestinal disease (P < 0.01). Weight loss was neither inevitable nor unremitting. Periods of weight stability (> 4 mo) occurred in 13 individuals (43%); 35 episodes of weight gain were identified, mostly related to recovery from opportunistic infection. These findings have important implications for our understanding of the natural history of weight loss in HIV infection.

Whole-body protein turnover from leucine kinetics and the response to nutrition in human immunodeficiency virus infection.

Whole-body protein metabolism was investigated in human immunodeficiency virus (HIV) infection by primed constant infusion of L-[1-13C]leucine in 8 control and 22 HIV-infected subjects (8 stage II; 14 stage IV disease), in postabsorptive and fed states. Postabsorptive leucine flux was increased 25% in subjects with stage IV HiV infection vs that in control subjects (130 +/- 13 vs 103 +/- 10 mumol leucine.kg-1.h-1, P < 0.001); both leucine disposal by protein synthesis (111.6 +/- 12.1 vs 82.3 +/- 9.2, P < 0.001) and release by protein degradation (129.7 +/- 13.1 vs 103.4 +/- 10.2, P < 0.001) were increased. No difference in leucine balance or oxidation was found but fat oxidation was greater in subjects with HIV infection (61.1 +/- 13.0% of energy) than in control subjects (47.6 +/- 13.7% of energy, P < 0.025). Stage II subjects had intermediate values of leucine flux, not significantly different from those of control subjects. Provision of parenteral nutrition for 4 h increased leucine flux with a switch in leucine balance from net loss to net gain; this response was quantitatively similar in all groups. HIV infection increases whole-body protein turnover but does not quantitatively impair the acute anabolic response to intravenous nutrition.

Production of beta-amyloid by primary human foetal mixed brain cell cultures and its modulation by exogenous soluble beta-amyloid.

Previous studies on beta-amyloid production have been carried out using transfected cells and cell lines. We measured the 40 and 42 amino acid forms of beta-amyloid released into the culture medium by primary human foetal mixed brain cell aggregate culture over 3 months. In this model, neurones and supporting cells are maintained in serum-free defined medium. The secretion of significant amounts of beta-amyloid 40 and 42 was observed throughout culture for three separate cultures. Levels of beta-amyloid 40 and 42 closely followed the neuronal content of the cultures as estimated by cellular neurone-specific enolase. Addition of synthetic beta-amyloid 1-40 to the cultures for 1 week at 35 days in vitro resulted in a dose-related reduction in cellular neurone-specific enolase levels. Primary human aggregate brain cell cultures produced multimeric beta-amyloid, as determined by immunoassay. beta-Amyloid-treated cultures released diminishing amounts of multimeric beta-amyloid and contained increasing amounts of intracellular multimeric beta-amyloid with increasing exogenous beta-amyloid. These results suggest that release of multimeric beta-amyloid into the extracellular environment by human primary neurones can be affected by the presence of extracellular beta-amyloid. This has implications for Alzheimer's disease in that beta-amyloid released into the extracellular environment by dead/dying neurones could modulate beta-amyloid release by surrounding neurones, potentially causing amplification of toxicity. Moreover, intracellular beta-amyloid oligomer-dependent neurotoxicity may be a component of neurodegeneration in Alzheimer's disease, and other conditions with increased beta-amyloid synthesis, suggesting anti-amyloid therapies for Alzheimer's disease may have to target intracellular beta-amyloid.

Depletion of neuroendocrine cells in rectal biopsy specimens from HIV positive patients.

AIMS: To compare the density of neuroendocrine cells in rectal biopsy specimens from human immunodeficiency virus (HIV) infected individuals with that of a control group. METHODS: Neuroendocrine cells in rectal biopsies were identified using an immunohistochemical stain for chromogranin and subsequently quantified using a method of linear intercept. RESULTS: Neuroendocrine cells were found to be significantly decreased in the HIV positive group. CONCLUSIONS: Loss of neuroendocrine cells may contribute to apoptotic bodies seen in this condition. This could be related to infection of these cells with HIV and could contribute to diarrhoeal disease in HIV infection.

Jejunal enteropathy associated with human immunodeficiency virus infection: quantitative histology.

Jejunal biopsy specimens from 20 human immunodeficiency virus (HIV) positive male homosexual patients were analysed and compared with those of a control group to determine whether the abnormalities were caused by the virus or by opportunistic infection. The degree of villous atrophy was estimated with a Weibel eyepiece graticule, and this correlated strongly with the degree of crypt hyperplasia, which was assessed by deriving the mean number of enterocytes in the crypts. The density of villous intraepithelial lymphocytes fell largely within the normal range, either when expressed in relation to the number of villous enterocytes or in relation to the length of muscularis mucosae. Villous enterocytes showed mild non-specific abnormalities. Pathogens were sought in biopsy sections and in faeces. Crypt hyperplastic villous atrophy occurred at all clinical stages of HIV disease and in the absence of detectable enteropathogens. An analogy was drawn between HIV enteropathy and the small bowel changes seen in experimental graft-versus-host disease. It is suggested that the pathogenesis of villous atrophy is similar in the two states, the damage to the jejunal mucosa in HIV enteropathy being inflicted by an immune reaction mounted in the lamina propria against cells infected with HIV.