Immune functions of the garment workers.

BACKGROUND: Occupational exposure to cotton dust, fibers, metal fumes and different chemicals used in the aparrel manufacturing industries cause a wide range of physical and psychological health problems in the garment workers that may also affect their immune function. OBJECTIVE: To assess the immune system function in garment workers. METHODS: A total of 45 workers of a garment factory, and 41 control subjects, not exposed to the garment working environment were enrolled in this study. In the study subjects, the complement system function was assessed as bactericidal activity on Escherichia coli DH5α cells using the standard plate count method. Serum complement components C3 and C4 were measured by immunoprecipitation, and IgG was measured by immunonephelometry. RESULTS: The bactericidal activity of serum complement in the garment workers (range: 93.5%-99.9%) was significantly (p<0.01) lower than that in the controls (range: 98.6%-100%). The heat-inactivated serum of the workers showed a significantly enhanced bactericidal activity. In the garment workers, the mean levels of complement C3, and C4 were 1.75 and 0.26 g/L, respectively that were close to those of the controls. The mean IgG level in the garment workers was 13.5 g/L that was significantly (p<0.001) higher than that in the controls. CONCLUSION: Working in a garment factory may affect the immune system.

Chronic carriers of hepatitis B virus in Bangladesh: a comparative analysis of HBV-DNA, HBeAg/anti-HBe, and liver function tests.

Serological markers of hepatitis B virus (HBV), liver function tests and quantitative estimation of HBV-DNA are important in the assessment of the state of infection and prognosis following treatment for hepatitis B. This study aimed to determine whether low-cost assays, eg hepatitis B e antigen (HBeAg) and liver function tests, could be used for the assessment of infectivity as an alternative to HBV-DNA estimation. We tested 125 hepatitis B carriers for HBeAg, antibody to HBeAg (anti-HBe), and serum HBV-DNA; we also carried out a range of standard liver function tests. Seventy-three subjects were positive and 52 were negative for HBeAg. Of the HBeAg positive cases, 3 were also positive for anti-HBe; of the HBeAg negative cases, 5 were also negative for anti-HBe. Of these 8 cases, 7 had no detectable HBV-DNA. Most of the HBeAg positive but anti-HBe negative subjects were positive for HBV-DNA (74.3%; 52/ 70) whereas most of the HBeAg negative and anti-HBe positive subjects (93.6%; 44/47) were also negative for HBV-DNA. Of 56 HBV-DNA positive individuals, alanine transaminase (ALT) was found to be raised in 69.6% (p=0.066) and aspartate transaminase (AST) was raised in 66.1% (p=0.011), while 67.9% had normal alkaline phosphatase (ALP) (p=0.054). HBeAg (p=0.018) and raised ALT (p=0.008) were found to be independent predictors for HBV-DNA positivity among HBV carriers. This study suggests that HBeAg positive and anti-HBe negative hepatitis B carriers with raised ALT and AST are likely to be positive for HBV-DNA; the combination of routine serology and biochemical tests may be considered as an alternative to HBV-DNA in evaluating the state of chronic HBV infection. However, HBV-DNA should be specifically assessed if discordance is observed between seromarkers and transaminases.

Immune response of Bangladeshi children with acute diarrhea who subsequently have persistent diarrhea.

BACKGROUND: Because altered immune responses may be a risk factor for persistent diarrhea, various aspects of the immune response were examined to elucidate the underlying immune mechanisms that may be involved in the development of persistent diarrhea. METHODS: Children (7-12 months of age) with watery diarrhea for 6 to 8 days from the Dhaka Hospital of the International Centre for Diarrheal Disease Research, Bangladesh (ICDDR,B), were enrolled. Children were classified as having acute diarrhea (AD) or persistent diarrhea (PD) if diarrhea resolved within 14 days or persisted for more than 14 days, respectively. Uninfected control children (n = 13), from the Nutrition Follow-Up Unit of ICDDR,B were also enrolled. Of the 123 children with diarrhea who were enrolled, 85 had AD and 38 had PD. Comparisons were performed for clinical features, nutritional status (weight for age, plasma transferrin, and serum albumin levels), and immune responses: neutrophil function; peripheral blood mononuclear cell function, delayed-type hypersensitivity (DTH) responses, plasma levels of immunoglobulins, tumor necrosis factor-alpha, and interferon-gamma. Univariate analyses were conducted to assess differences among the three groups of children and between children with AD and PD. Logistic regression was performed to determine risk factors for PD. RESULTS: There were no differences in clinical features and nutritional status among the groups of children studied. More children in whom PD developed had a negative DTH response to tuberculin than those with AD (P = 0.021). Also, a negative DTH response to tuberculin was a significant risk factor for PD (odds ratio [OR] = 3.8, 95% confidence interval [CI] = 1.5-9.9). CONCLUSIONS: Children with acute diarrhea with a negative DTH response to tuberculin are more likely to have development of persistent diarrhea.

Peripheral blood neutrophil responses in children with shigellosis.

Alterations in peripheral blood neutrophil function are known to occur in patients with colitis and may have a role in precipitating nonspecific tissue injury. It is not known whether neutrophil function is altered in patients with Shigella dysenteriae type 1 infection, during which there is extensive colitis and which may be associated with life-threatening complications in young children. Three aspects of peripheral blood neutrophil function, polarization, attachment to yeast particles, and locomotion, were therefore studied in 111 children with S. dysenteriae type 1 infection and 57 children without any infection. All children were aged 12 to 60 months. Of the children with S. dysenteriae type 1 infection, 42 had leukemoid reaction, hemolytic-uremic syndrome, or septicemia (complicated shigellosis), while the others did not (uncomplicated shigellosis). Polarization and locomotion in the absence of chemoattractants and in response to N-formylmethionyl-leucylphenylalanine (FMLP) and the lipopolysaccharide (LPS) of S. dysenteriae type 1 were determined. Attachment to unopsonized and opsonized yeast particles was also determined. Children with shigellosis (uncomplicated or complicated) had more polarized neutrophils with and without chemoattractants than uninfected children (P < 0.05). Children with complicated shigellosis had more polarized neutrophils with FMLP at 10(-7) and 10(-6) M (P < 0.05) and with LPS than children with uncomplicated shigellosis (P < 0.05). At 3 to 5 days after enrollment, the numbers of polarized neutrophils with 10(-8), 10(-6), and 10(-5) M FMLP declined in children with uncomplicated shigellosis but not in those with complicated shigellosis. Attachment to yeast particles was similar in all three groups of children. Locomotion was inhibited by LPS in children with shigellosis (P < 0.05), whether it was uncomplicated or complicated, compared with locomotion in uninfected children. Finally, neutrophil polarization in uninfected children was negatively influenced by nutritional status. Thus, poorly nourished uninfected children had more polarized neutrophils with FMLP at 10(-9) M (P < = 0.02) and 10(-5) M (P = 0.043) than their better-nourished counterparts. In summary, altered neutrophil responses are associated with both uncomplicated and complicated shigellosis.

Peripheral blood granulocytes and mononuclear cell responses in monkeys with experimental shigellosis.

Changes in neutrophil response to N-formyl-methionyl-leucyl-phenylalanine (FMLP) and the phenotype of peripheral blood mononuclear cells were studied in monkeys after oral challenge with Shigellae. Monkeys were first challenged with S. dysenteriae 1 which caused shigellosis in some of the monkeys. After recovery, the monkeys were rechallenged with S. flexneri 2a. No difference in sensitivity was observed in the monkeys during shigellosis caused by either S. dysenteriae 1 or S. flexneri 2a. The optimal dose of FMLP for neutrophil polarization, a measure of early cell activation, in normal healthy monkeys was 10(-7) M when 67% of the neutrophils were polarized. Neutrophils from monkeys ill with shigellosis required higher doses of FMLP (10(-6) and 5 x 10(-7) M) for maximum polarization. As the monkeys recovered, a gradual decrease in the doses of FMLP for optimal neutrophil polarization was also observed. The percentage of CD2-positive T lymphocytes, the earliest marker for T lymphocytes in the peripheral blood, decreased when the monkeys developed shigellosis and returned to normal levels as the monkeys improved. However, there was no change in the percentage of CD20-positive peripheral blood B lymphocytes.

Restoration with phorbol ester of a locomotor defect in human leukaemic lymphocytes: a visual study of chronic lymphocytic leukaemia cells.

Visual assays were used to study the effect of phorbol myristate acetate (PMA) on the locomotion of lymphocytes from 14 patients with chronic lymphocytic leukaemia (CLL). Previous reports had shown that CLL cells from blood were defective in locomotor capacity and that adding PMA to the cells did not restore locomotion in a short-term (30 min) assay, but did stimulate unusual non-locomotor, multipolar, morphologies in a small proportion of the cells. Here we describe experiments in which CLL cells were cultured for 48 h in PMA. Many of the cells acquired locomotor morphologies with front-tail polarity which was unlike the short-term multipolar morphology. These cultured cells were also capable of locomotion and invaded collagen gels. Autoradiography suggested that after culture in PMA, the locomotory cells were the most active cells in 3H-uridine uptake. A computer analysis suggested that the cells in locomotor morphology were the same cells that increased in size. These findings suggest that, to acquire locomotor capacity, CLL lymphocytes require an appropriate signal to allow the cells to enter the cell cycle. During G1 the lymphocytes develop the capacity for locomotion. Long-term, but not short-term, culture in PMA provides such a signal but the mechanism by which it does so is unclear.

Evaluation of methods for isolating human peripheral blood monocytes. Studies on chemotactic locomotion and other functional characteristics.

We have compared six different methods of purifying human blood monocytes for their usefulness in relation to assays of polarization, locomotion and chemotaxis. For polarization assays it is essential to prepare an unstimulated, spherical, cell population in suspension. The techniques compared were based either on density differences between monocytes and lymphocytes using Percoll or Nycodenz, or on the separation of adherent monocytes from non-adherent cells on protein-coated surfaces, i.e., foetal calf serum (FCS); gelatin-FCS; gelatin-plasma; baby hamster kidney (BHK) microexudate coats. The BHK microexudate technique (Ackerman and Douglas, 1978) gave the best yield and purity of monocytes. These were spherical and had not been activated by the separation procedure. This technique provided monocytes in suspension that were functionally normal in locomotion and chemotaxis assays, phagocytosis, chemiluminescence and Fc receptor expression. To achieve a good yield of spherical cells, it was necessary to use tubes to which monocytes did not adhere. Siliconized glass was superior to tissue culture plastic for this purpose.

Recombinant IL-4 and IFN-gamma activate locomotor capacity in human B lymphocytes.

Recombinant human interleukin-4 (IL-4) or interferon-gamma (IFN-gamma) were added to cultures of B-enriched human lymphocytes from normal blood, or to the lymphocytes from five patients with chronic lymphocytic leukaemia (CLL). IL-4 and IFN-gamma caused the B lymphocytes to acquire locomotor capacity, as judged by morphological polarization and invasion of collagen gels. This was detectable in normal B cells within a few hours of culture and fully developed by 24-48 hr. It was inhibited by the presence of cyclosporin A. The responding, motile cells also increased in size. These findings suggest that B lymphocytes acquire locomotor capacity early in growth, as the cells move from G0 to G1. IL-4 or IFN-gamma had no direct effect in polarizing lymphocytes in a short-term (30-min) assay, thus they do not behave like chemotactic factors. They slowly increase the proportion of locomotor cells in B-lymphocyte populations, and these motile cells respond by polarization to factors released by the growing cells into the medium.

Effects of phorbol esters on shape and locomotion of human blood lymphocytes.

The effects of phorbol esters on shape change and locomotion of human blood lymphocytes were studied both immediately after separating the cells from blood and after overnight culture. Phorbol myristate acetate (PMA), phorbol dibutyrate (PDB) and related esters produced complex shape changes in lymphocytes at both times. These shapes were analysed quantitatively using objective measurements derived from the moments of cell shapes. Immediately after removal from blood, many lymphocytes (20-60% of the total) protruded and retracted veils or spikes at more than one point on the cell surface. The morphology of these cells was not typical of locomotor cells. Usually, formation of a veil was not followed by a contraction wave moving down the cell, though some cells did show contraction waves, and some moved into collagen gels or filters. After overnight culture, a high proportion (70-80%) of cells had changed shape in PMA and PDB. Although the shapes were still atypical, they resembled classical locomotor morphology more closely; veils formed at one point on the cell surface tended to persist, and contraction waves and constriction rings were seen in many cells. These cells moved in large numbers into collagen gels or filters. Comparison of the paths traversed by PMA-treated lymphocytes in collagen gels suggested that cells cultured in PMA for 24 h pursued more persistent paths that those in short-term culture, but the difference was not marked. We suggest that phorbol esters induce immediate shape change without inducing the complete sequence of motor events necessary for efficient locomotion, whereas after prolonged culture in phorbol esters, locomotion is more efficient, possibly because phorbol esters, like other growth activators, stimulate events during the G1 phase of growth that are necessary for full expression of locomotor capacity.

Chemotactic factor-induced polarization, receptor redistribution, and locomotion of human blood monocytes.

The locomotor response of human blood monocytes to chemotactic factors was studied using a polarization assay on cells in suspension and by filming locomotion on albumin-coated glass. Cells in optimal (5 x 10(-9) M) but uniform concentrations of N-formyl-methionyl-leucyl-phenylalanine (FMLP) polarized well and showed a 'persistent random walk' type of locomotion, whereas in supraoptimal concentrations (10(-7) M), the cells took erratic paths and polarized poorly, suggesting that monocytes cannot develop an anteroposterior polarity if hit by ligand molecules at many points on the cell surface simultaneously. Monocyte polarization in chemotactic factors at 37 degrees was transient and was gradually lost after 15-20 min. Likewise, the ability to form Fc rosettes after this time was gradually lost, suggesting loss of functional receptors from the cell surface with time. In optimally polarized cells, Fc rosettes were frequently localized at the head of the cell. This localization also was lost with time. Using pure chemotactic factors (FMLP, C5a, leukotriene B4) we found, as reported earlier (Cianciolo & Snyderman 1981), that polarization was restricted to a subpopulation (approximately 60% of cells) that responded to multiple attractants. However, 80-90% of monocytes polarized in response to combinations of any of the above pure attractants with candida-activated serum. This suggests that the subpopulation that lacks receptors for classical chemotactic factors nevertheless has locomotor capacity and can respond to undefined factors in activated serum, and that the great majority of blood monocytes is motile if appropriately stimulated.

The defect of lymphocyte locomotion in chronic lymphocytic leukaemia: studies of polarization and growth-dependent locomotion.

This paper reports a study of the locomotor behaviour of the lymphocytes from 17 patients with chronic lymphocytic leukaemia (CLL). The cells were studied both immediately after separation from blood and after culture for 24 to 48 h with a range of growth activators. Cells direct from blood were tested for polarization in fetal calf serum (FCS 20%), phorbol myristate acetate (PMA 10(-7)M) and colchicine (10(-5)M). The polarization of lymphocytes from CLL patients with high white cell (WBC) counts (greater than 10 X 10(9)/litre) was very poor in FCS and PMA, though the cells from about half of these patients responded well to colchicine. The response of cells from patients with low white cell counts was the same as that of controls. The growth activators, PHA (1 micrograms/ml), anti-CD3 antibody (OKT3 2.5 ng/ml), Cowan strain Staphylococcus aureus (SAC; 1.5 X 10(7)/ml) and PMA (10(-8)M) induced an increase in the proportion of locomotor lymphocytes from controls and from CLL patients with low white cell counts during 24 h of culture. Cells from patients with high white cell counts showed very little increase in locomotor forms in any activator including PMA and the B cell mitogen SAC. This defect was seen in both polarization assays and collagen gel invasion assays. The findings suggest that CLL lymphocytes have a defect of locomotion demonstrable at two levels: (a) the cells fail to respond by polarizing immediately upon stimulation; colchicine treatment reverses this defect in some cases; (b) they also fail to acquire locomotor capacity during culture with activators of growth.

The use of collagen or fibrin gels for the assay of human neutrophil chemotaxis.

Neutrophil leucocytes are known to migrate actively into 3-dimensional gels of collagen or fibrin. In this paper, we have used such gels to study chemotaxis of human blood neutrophils towards gradient sources of formyl-methionyl-leucyl-phenylalanine (FMLP) using 2 assay systems. The first resembled the micropore filter assay in that neutrophils on the upper surface of collagen gels were allowed to invade in the presence of either an isotropic concentration or a gradient of FMLP. Neutrophils invaded the gel vigorously in both cases. The effect of the gradient was assessed by determining the population distribution at different levels in the gel. Cells moving randomly should be distributed normally, and directional locomotion should cause deviation from normal distribution. Such a deviation was seen, but was of marginal significance. A more direct demonstration of chemotaxis was achieved by the second assay in which an agarose slab containing FMLP was incorporated into a gel, and the paths of nearby neutrophils were filmed. These cells showed an unequivocal directional response to the FMLP gradient. Protein gels can thus be used in the same way as both the presently used filter assays and visual assays using plane substrata, but with the advantage of providing a more physiological environment for the study of chemotaxis than either.