Effect of the metabolic syndrome on male reproductive function: a case-controlled pilot study.

The metabolic syndrome (MetS) is a constellation of various risk factors. This study aimed to investigate the effect of MetS on testosterone and progesterone, and semen parameters, in a case-controlled pilot study. Male patients (n = 54) had body mass index, waist-to-hip ratio (WHR) and blood pressure recorded. Blood was analysed for HDL cholesterol, triglycerides and glucose. Saliva was assayed for free testosterone and free progesterone. Ejaculates were analysed for volume, sperm concentration, total sperm count, motility, vitality, mitochondrial membrane potential (MMP), DNA fragmentation and leucocyte concentration. Participants were divided into the control group (n = 28) and the MetS group (n = 26). Differences were found between the groups for body mass index, WHR, blood pressure, high-density lipoprotein (HDL), triglycerides and glucose. The MetS group showed significant reductions in sperm concentration (P = 0.0026), total sperm count (P = 0.0034), total motility (P = 0.0291), sperm vitality (P = 0.002), MMP (P = 0.0039), free testosterone (P = 0.0093) and free progesterone (P = 0.0130), while values for DNA fragmentation increased (P = 0.0287). Results indicate that patients with MetS have compromised sperm parameters in the absence of leucocytospermia. A reduction in free progesterone suggests that steroidogenesis cascades may be compromised. It is hypothesised that a systemic pro-inflammatory state with oxidative stress associated with MetS may provide a novel explanation.

TUNEL assay and SCSA determine different aspects of sperm DNA damage.

For the determination of sperm DNA damage, different assays are used. However, no further distinction is made and the literature generally speaks about DNA damage. Thus, this study aimed at comparing the sperm chromatin structure assay (SCSA) and the TUNEL assay. In 79 patients, sperm DNA damage was determined flow cytometrically using the SCSA and the TUNEL assay. Moreover, normal sperm morphology was evaluated according to strict criteria. A statistical comparison of the two methods was performed using standard correlations, Bland and Altman plots, Passing-Bablok regressions and concordance correlation. Results show a significant difference between P- and G-pattern morphology only for the mean channel fluorescence of the SCSA. Spearman's rank correlations between the different parameters of both assays, SCSA and TUNEL, revealed significant associations between the parameters of the assays. However, when applying Bland and Altman plots, Passing-Bablok regression and concordance correlation results showed that these methods are not comparable. These different techniques determine different aspects of sperm DNA damage, i.e. 'real' DNA damage for the TUNEL assay and 'potential' DNA damage in terms of susceptibility to DNA denaturation for the SCSA. Thus, one should clearly distinguish between the different assays, not only practically and methodologically but also linguistically.

Complexes of alpha 1-microglobulin and monomeric IgA in multiple myeloma and normal human sera.

alpha 1-Microglobulin (alpha lm), a glycoprotein heterogeneous in charge, was reported to occur both as a 31-kilodalton (kd) monomer [low mol. wt alpha lm (LMW-alpha lm)] and as polymers or complexes formed with other plasma proteins including IgA [high mol. wt alpha lm (HMW-alpha lm)]. The present study was designed to characterize HMW-alpha lm in normal human serum and in myeloma sera. The following sera were selected: five IgG, 16 IgA and four Bence-Jones protein myelomas. alpha lm was identified by specific monoclonal antibodies in competitive radioimmunoassay and solid-phase ELISA. HMW-alpha lm was found to be associated almost exclusively with monomeric IgA and possibly in very small proportion with dimeric IgA. Ever in cases of predominantly dimeric IgA myelomas, alpha lm was associated with the monomeric form of the monoclonal IgA. The molar ratio of HMW-alpha lm to monomeric IgA never exceeded 3.5% and it was estimated to range from 0.5 to 1.4% in normal serum. No association with other proteins than IgA and no alpha lm polymers were found in IgA myeloma. Two types of HMW-alpha lm-IgA complexes were found: (a) those that were dissociable into IgA and LMW-alpha lm after mild reduction, and (b) those which were dissociated only after complete reduction of the complexes into IgA and an 88-90-kd component bearing alpha lm but no IgA epitopes. It was concluded that either of the two molecular species of alpha lm bearing common epitopes, with apparent mol. wts of 31,000 and 88,000-90,000, respectively, could form stable complexes with monomeric IgA. The association is likely to be performed through disulfide bridges. Nearly all the 88-90-kd but only a small proportion of the 31-kd component is associated with IgA.

Immunohistological localization of alpha 1-microglobulin in normal rat tissues.

The tissue distribution of rat alpha 1-microglobulin (alpha 1-m) was studied by indirect immunofluorescence in various rat tissues using a polyvalent rabbit antiserum to the purified antigen and a monoclonal antibody (H23) to the human homologue, in parallel with a polyclonal anti-rat IgA antiserum. It was found that all tissues stained by anti-IgA were also alpha 1-m positive; these tissues included tissues of the stomach, duodenum, ileum, colon, pancreas, trachea, esophagus and jejunum. However, the observation that IgA plasma cells as well as secretory cells, while positively stained by anti-IgA, are alpha 1-m negative suggests that the association between IgA and alpha 1-m occurs at a postsecretory stage, after the IgA molecules have been transported across the epithelial cells. Additionally, hepatocytes were intensely stained by anti-alpha 1-m antibodies, indicating that the liver, as already suggested by metabolic studies on isolated guinea-pig liver explants, may be responsible for the synthesis of this protein. Among lymphoid tissues, an intense and homogeneous staining was observed in the thymus and the white pulp of the spleen. Sections of lymph nodes, however, showed differential staining; apart from a few isolated dendritic cells in the mantle region of the lymphoid follicles, the germinal centers and medullary cords showed no staining with anti-alpha 1-m antibodies. The paracortical cells, macrophages in the subcapsular sinus, and interfollicular lymphocytes showed intense cytoplasmic staining with anti-alpha 1-m antibodies. In other tissues, macrophages, monocytes, tissue histiocytes, and dendritic cells were alpha 1-m positive. Although they confirm the presence of alpha 1-m in the lymphoid tissues, as already reported in man, these results show that the protein is also present in hepatocytes and in exocrine fluids containing IgA. Since alpha 1-m, like secretory component, can bind to IgA to form stable complexes, these two heavily glycosylated proteins may have similar biologic properties.

Endocytosis of class II histocompatibility antigens and formation of intracytoplasmic granules at the final differentiation stage of human B lymphocytes.

The expression of histocompatibility antigens was investigated using several human lymphoid cell lines representative of different maturation stages of the B-cell lineage. Class II HLA antigens were found at the surface of all cell lines. However, in the myeloma cell line U266, an intracellular macrovesicular pool of these antigens was found in some cells. It originated from microvesicular endocytosis of the surface antigen, subsequently leading to cells bearing HLA class I but not class II antigens. Since the latter play a major role in cellular interactions regulating B-cell differentiation, this phenomenon may be linked to the final stage of maturation of B lymphocytes into plasma cells.

A randomised placebo-controlled trial of the efficacy of beta-sitosterol and its glucoside as adjuvants in the treatment of pulmonary tuberculosis.

OBJECTIVE: To evaluate the adjuvant effect of beta-sitosterol and its glucoside in the treatment of culture proven pulmonary tuberculosis (PTB). DESIGN: A blinded randomised placebo-controlled trial in culture proven drug sensitive PTB. Patients were hospitalised for the duration of treatment and evaluated at monthly intervals with regard to sputum culture positivity, chest radiography, weight gain, Mantoux test response, routine haematology and liver functions. STATISTICAL EVALUATION: General linear models for repeated measures (SAS GLM package) compared the interaction effects, group effects and time effects of findings in 19 patients receiving sitosterols with those in 18 patients receiving a placebo (talcum powder). Absolute values and change from baseline values were evaluated, although only the latter are reported. RESULTS: Weight gain was significantly greater in the sitosterol group (mean weight gain 8.9 kg) than the placebo group (mean gain 6.1 kg) (P = 0.0023 group effects; P = 0.0001 for time effects). Speed of achieving culture negativity, radiological improvement and induration on Mantoux testing was similar in the two groups. Change in lymphocyte counts from baseline was significantly higher in the sitosterol group (P = 0.0001 and P = 0.0001 for group and time effects) as was the increase in eosinophil counts (P = 0.0001 and P = 0.0137 for group and time effects). CONCLUSION: The study has shown significantly improved weight gain and higher lymphocyte and eosinophil counts in PTB patients receiving sitosterols in addition to an efficacious antituberculosis regimen. Sitosterols and their possible mode of action should now be evaluated in larger numbers of tuberculosis patients and in diseases with a similar immunopathogenesis.

Plant sterols and sterolins: a review of their immune-modulating properties.

Beta-sitosterol (BSS) and its glycoside (BSSG) are sterol molecules which are synthesized by plants. When humans eat plant foods phytosterols are ingested, and are found in the serum and tissues of healthy individuals, but at concentrations orders of magnitude lower than endogenous cholesterol. Epidemiological studies have correlated a reduced risk of numerous diseases with a diet high in fruits and vegetables, and have concluded that specific molecules, including b-carotene, tocopherols, vitamin C, and flavonoids, confer some of this protective benefit. However, these epidemiologic studies have not examined the potential effect that phytosterols ingested with fruits and vegetables might have on disease risk reduction. In animals, BSS and BSSG have been shown to exhibit anti-inflammatory, anti-neoplastic, anti-pyretic, and immune-modulating activity. A proprietary BSS:BSSG mixture has demonstrated promising results in a number of studies, including in vitro studies, animal models, and human clinical trials. This phytosterol complex seems to target specific T-helper lymphocytes, the Th1 and Th2 cells, helping normalize their functioning and resulting in improved T-lymphocyte and natural killer cell activity. A dampening effect on overactive antibody responses has also been seen, as well as normalization of the DHEA:cortisol ratio. The re-establishment of these immune parameters may be of help in numerous disease processes relating to chronic immune-mediated abnormalities, including chronic viral infections, tuberculosis, rheumatoid arthritis, allergies, cancer, and auto-immune diseases.

Clinical outcome of HIV positive patients with moderate to severe burns.

Human immunodeficiency virus (HIV) infection is a world wide and growing problem. Little is found in the literature concerning the treatment and outcome of patients suffering from HIV infection who are treated for burns. The aim of this study was to assess whether the outcome of HIV positive patients suffering from burn wounds differed from those who do not have HIV infection. Thirty three patients formed the HIV positive study group. HIV negative controls were matched for age, degree of burns, sex and inhalation injury. The mean age of the patients was 31.6 years and the mean total body surface burn was 26.4%. There was no significant difference in the outcome of the two groups in terms of mortality or treatment parameters measured. Two patients had stigmata of AIDS (tuberculosis) and both died. One patient, with a CD4 count of 228, developed severe necrotizing fasciitis. In keeping with other studies looking at the outcome of HIV positive patients in an Intensive Care Unit setting, we concluded that a HIV positive patient, who suffers from a burn wound and has no stigmata of AIDS, should be treated similarly to a HIV negative patient.

Immune deficiency in familial duodenal atresia.

The familial occurrence of duodenal atresia is uncommon. This study evaluated the inheritance patterns, the nature and associations, and the presence of immunologic deficits in duodenal atresia recurring in at least three siblings each in two nonrelated families. In the first family, an association with Fanconi's anemia was observed in three of seven pregnancies (2 boys, 1 girl) suggesting an autosomal recessive mode of transmission. Patients died as a result of overwhelming (fungal) septicemia in association with pancytopenia. In a second family, identical multiple atresias occurred in two female siblings born 18 months apart and a third child with a duodenal stenosis. Overwhelming sepsis and a T-cell dysfunction was seen in the postoperative period, which had partially corrected by follow-up at 5 months. A history of family occurrence of duodenal atresia should alert the physician to the possibility of associated pathology including immune deficiency states.

Interleukin-1α Induction in Human Keratinocytes (HaCaT): An In Vitro Model for Chemoprevention in Skin.

Long-term exposure to UV irradiation and toxic chemicals is associated with chronic inflammation that contributes to skin cancer development with interleukin-1 alpha (IL-1α), constitutively produced by keratinocytes, playing a pivotal role in skin inflammation. The aim of this study was to investigate the modulation of IL-1α production in the HaCaT keratinocyte cell line. Phorbol 12-myristate 13-acetate failed to induce IL-1α in HaCaT cells, and this might be associated with the specific deficiency known to affect downstream signalling of the MEK/ERK pathway in these cells. The calcium ionophore, ionomycin, slightly enhanced the production of intracellular (icIL-1α), but this resulted in a necrotic release at higher concentrations. UV-B exposure significantly increased the production of icIL-1α in a dose-dependent manner with a maximal induction exhibited at 24 h with minimal necrotic and apoptotic effects. Validation of the HaCaT cell model indicated that the nonsteroidal anti-inflammatory drug (NSAID), ibuprofen, and the glucocorticoid, dexamethasone, inhibited icIL-1α production, and this was associated with a slight inhibition of cell viability. The UV-B-induced keratinocyte cell model provides an in vitro system that could, apart from phorbol ester-like compounds, be utilised as a screening assay in identifying skin irritants and/or therapeutic topical agents via the modulation of IL-1α production.

Plasma sarcosine does not distinguish early and advanced stages of prostate cancer.

INTRODUCTION: Diagnosis of prostate cancer by prostate specific antigen (PSA) is error-prone and cannot distinguish benign prostatic hyperplasia (BPH) from malignant disease, nor identify aggressive and indolent types. METHODS: We determined serum sarcosine (N-methylglycine) in 328 cancer patients by gas chromatography (GC)/mass spectroscopy (MS) and searched for correlations with early (stage T1/T2) and advanced (stage T3/T4) disease. RESULTS: Serum sarcosine of male control patients ranged from 1.7 µmol/l to 4.8 µmol/l. In prostate cancer patients, sarcosine ranged from 2.8 µmol/l to 20.1 µmol/l. Expressed as the sarcosine/alanine ratio, serum control values were 9.4 ± 5.5 x 10(-3) (mean ± SD) compared with 21.6 ± 9.0; 28.5 ± 16.6; 22.7 ± 7.7 and 22.2 ± 11.0 for patients diagnosed with T1, T2, T3 and T4 prostate tumours, respectively. The small differences between T1, T2, T3 and T4 patients were not statistically significant (p=0.51). However, the conventional PSA marker significantly correlated with T stage in these patients (r=0.63; p<0.009). CONCLUSIONS: The median sarcosine/alanine ratios among patients with early and advanced prostatic cancer ranged from 21.6 ± 9.0 to 28.5 ± 16.6 and were fairly constant, showing no statistically significant differences between T-stages. The results are consistent with published data in urine and serum which find differences between controls and patients with metastatic prostate cancer to be small and sarcosine to be uninformative regarding prostate cancer progression. By multi-comparison of PSA with T-stages in the same group of patients, we found significant correlations confirming the well-known merits and limitations of this marker.

The role of phytosterols and phytosterolins in immune modulation: a review of the past 10 years.

Although plant sterols (phytosterols) were chemically described in 1922, their biological role in human and animal health has been underestimated. Their ability to control cholesterol plasma levels in hypercholesterolimic patients was first described in 1983 when the structure of phytosterols implied that they could, by steric hindrance, inhibit the absorption of cholesterol from our diets. This has led to the development of functional foods containing high contents of these plant molecules or their esters as cholesterol controlling foods. Over the last 15 years, however, several reports have appeared in the literature indicating that phytosterols have some immunological activity as highlighted in animal models of inflammation or even in in-vitro and in-vivo models of cancer (colorectal and breast cancer). These findings were paralleled by epidemiological studies correlating the reduced risk of numerous diseases and the dietary intake of phytosterols. It is only in the last 10 years, however, that their direct immune modulatory activity on human lymphocytes has been proven and the mechanism of action in cancer cells has been elucidated. The use of phytosterols as supportive therapies in certain chronic conditions has been tested under clinical trial conditions. This review presents a summary of the in-vitro and in-vivo studies published to date.

The detection of pyrogens in sera from patients with symptoms of sepsis using an ex vivo whole blood culture assay.

The aim of this study was to investigate whether the ex vivo whole blood culture (WBC) assay system can be used to detect pyrogens in blood from patients with symptoms of sepsis. Blood samples from 35 patients with symptoms of sepsis were assayed for bacterial contamination using the radiometric blood culture assay. Serum from the same patients were screened for IL-6, C-reactive protein (CRP) and pyrogens using the whole blood culture assay. Serum samples from 26 patients tested positive for pyrogens. Of the 26 patients with pyrogenic serum, 15 had elevated serum IL-6 levels and 19 had elevated CRP levels. Only two of the samples had positive blood cultures as detected by the routine radiometric assay. Both of these patients had high serum CRP and pyrogen levels, while only one of them had an elevated serum IL-6 level. These results show that the WBC is very sensitive in detecting pyrogens in serum of patients. This technique can be a useful tool to quantitate pyrogens in sera from patients with symptoms of sepsis and to determine whether their clinical symptoms are caused by pyretic substances in their circulatory system.

IL-6 secretion by ex vivo whole blood cultures upon allergen stimulation.

Allergen induced IL-6 synthesis by whole blood cultures was compared with skin prick allergen test results for the same group of individuals. Whole blood cultures from both allergic and non-allergic individuals secrete IL-6 at high allergen concentrations. When whole blood cultures from controls were incubated with serial dilutions of allergens it was found that IL-6 induction was abolished at lower allergen dilutions (allergen threshold concentration or ATC). When whole blood cultures from patients with allergic rhinitis were stimulated with ATC it was found that some allergens induced IL-6 secretion. The allergens inducing IL-6 and the level of IL-6 secreted were dependent on the patient. The induction of IL-6 secretion by the cultures at ATC correlated very significantly with the patient's skin prick test results (r = 0.71 1; p = 0.0003).

Flow cytometric analysis of the Th1-Th2 balance in healthy individuals and patients infected with the human immunodeficiency virus (HIV) receiving a plant sterol/sterolin mixture.

The Th1--Th2 balance plays a pivotal role in determining the outcome of an immune response to an infectious organism. It is proposed that during HIV infection, disease progression is characterized by a loss of Th1 activity, a shift to a more 'allergic' Th2-type response and hence loss of cytotoxic cell activity against infected host cells. This study was undertaken to investigate this balance in three groups of individuals: HIV-negative volunteers (n=10), a group of HIV-infected patients on no therapy (n=10) as well as a group of patients managed with a mixture of plant sterols/sterolins (n=9). In parallel, their response to mitogens and the subsequent expression of the activation antigen CD69 was measured. This study was conducted by three-colour flow cytometry in order to obviate the less sensitive cytokine secretion assays that have yielded controversial results. The results indicate that HIV-infected patients on no therapy exhibit a pre-dominant Th2 response (IL-4 secretion), whereas those on the sterol/sterolin mixture exhibit a beneficial Th1 response (IFN-gamma). Surprisingly, in both patient groups, the expression of CD69 was abnormally low when compared to the uninfected volunteers, implying that chronic activation is already present in vivo. It appears that the detrimental Th2 driven response might be swung to the more beneficial Th1 response with the immune modulatory sterols/sterolin mixture. Clinical use of this mixture in HIV infection has yielded results which corroborate the above observations in that patients using the plant sterol/sterolin mixture maintain their CD4 cell numbers over an extended period of time in the absence of any anti-retroviral therapy.

Major histocompatibility complex class II invariant chain expression in non-antigen-presenting cells.

In contrast to the generally accepted belief, the major histocompatibility complex (MHC) class II invariant chain (Ii) is commonly expressed intracellularly in cells that do not present exogenous antigens. Such cells include resting peripheral blood T cells and natural killer (NK) cells. In T cells, the Ii is associated with a 77 000 molecular-weight molecule (p77) that has yet to be identified. This molecule is co-precipitated with the anti-Ii monoclonal antibody (mAb) VCD-1, but not with mAb BU-45. This suggests that in the p77-Ii complex, the extracellular epitope of Ii recognized by BU-45 is hidden, whereas the Ii epitope for VCD-1 remains exposed. In antigen-presenting cells (APCs), p77 association with the Ii was minimal, if detectable. The p77-Ii association in non-professional APCs suggests that the Ii may have another, more general, function other than the one accepted in antigen presentation.

Characterization of rat alpha 1 microglobulin.

A molecule heterogeneous in charge with an apparent MW of 30 000 d and associated chromophore and carbohydrate was isolated from rat urine. When compared to human alpha 1-m its fluorescence spectra as well as staining pattern on agarose gel electrophoresis or electrofocussing were very similar. Furthermore, the specific antiserum prepared against the molecule detects an antigenic determinant also detected by monoclonal antibodies against human alpha 1-m. For these reasons this protein can be considered as the rat equivalent of human alpha 1-m.

Localization of alpha 1-microglobulin (HC protein) in normal human tissues: an immunohistochemical study using monoclonal antibodies.

Alpha-1-microglobulin is a low molecular weight (approximately 30 000 d) glycoprotein present in biological fluids. It is heterogeneous in charge. A monoclonal antibody was used to investigate the tissue distribution of the protein in normal human tissues and cell lines by indirect immunofluorescence and immunoperoxidase techniques. The protein was demonstrable in cells of the monocyte-macrophage lineage, in thymus and T cell dependent areas of spleen, lymph node and tonsils. It was detected in several lymphoid or nonlymphoid cell lines but not in peripheral blood lymphocytes. The microglobulin was also detectable in the cytoplasm of hepatocytes. Finally, it was observed in glandular secretions (sudoral glands and mucosal glands of the digestive tract) where it may be associated with IgA. Possible explanations for the highly divergent results previously reported with polyclonal antisera to alpha 1 microglobulin are discussed.

Peripheral blood lymphocyte response in patients with superficial transitional cell carcinoma of the bladder treated with intravesical Bacillus Calmette-Guérin--a useful marker of response?

Intravesical instillation of Bacillus Calmette-Guérin (BCG) offers safe and effective short-term/long-term treatment for superficial transitional cell carcinoma (TCC) and TCC in situ of the bladder. However, 17 to 42% of patients may experience recurrence in spite of this therapy and a marker of effective treatment is of paramount importance. In this study the in vitro response of peripheral blood lymphocytes (PBL) to BCG was analysed in 10 patients with superficial TCC and TCC in situ before and during BCG instillations. The in vitro response of PBL to BCG, expressed as a stimulatory index (SI), revealed that 6 patients had a SI > 5 and 4 patients had a SI < 5. None of the former patients had recurrence of TCC during a mean follow-up of 17 months, while all of the latter patients experienced recurrence of TCC within 9 months. It was concluded that the in vitro response of PBL to BCG may be used as a marker of response to intravesical BCG treatment.