Vitamin D and prostate cancer.

Our recent epidemiological study (Ahonen et al., Cancer Causes Control 11(2000) (847-852)) suggests that vitamin D deficiency may increase the risk of initiation and progression of prostate cancer. The nested case-control study was based on a 13-year follow-up of about 19000 middle-aged men free of clinically verified prostate cancer. More than one-half of the serum samples had 25OH-vitamin D (25-VD) levels below 50 nmol/l, suggesting VD deficiency. Prostate cancer risk was highest among the group of younger men (40-51 years) with low serum 25-VD, whereas low serum 25-VD appeared not to increase the risk of prostate cancer in older men (>51 years). This suggests that VD has a protective role against prostate cancer only before the andropause, when serum androgen concentrations are higher. The lowest 25-VD concentrations in the younger men were associated with more aggressive prostate cancer. Furthermore, the high 25-VD levels delayed the appearance of clinically verified prostate cancer by 1.8 years. Since these results suggest that vitamin D has a protective role against prostate cancer, we tried to determine whether full spectrum lighting (FSL) during working hours could increase serum 25-VD concentrations. After 1-month exposure, there was no significant increase in the serum 25-VD level, although there was a bias towards slightly increasing values in the test group as opposed to decreasing values in controls. There was no significant change in the skin urocanic acid production. The possibility to use FSL in cancer prevention is discussed. In order to clarify the mechanism of VD action on cell proliferation and differentiation, we performed studies with the rat and human prostates as well prostate cancer cell lines. It is possible that 25-VD may have a direct role in the host anticancer defence activity, but the metabolism of vitamin D in the prostate may also play an important role in its action. We raised antibodies against human 1alpha-hydroxylase and 24-hydroxylase. Our preliminary results suggest that vitamin D is actively metabolised in the prostate. Vitamin D appears to upregulate androgen receptor expression, whereas androgens seem to upregulate vitamin D receptor (VDR). This may at least partially explain the androgen dependence of VD action. VD alone or administered with androgen causes a suppression of epithelial cell proliferation. VD can activate mitogen-activated kinases, erk-1 and erk-2, within minutes and p38 within hours. Also, auto/paracrine regulation might be involved, since keratinocyte growth factor (mRNA and protein) was clearly induced by VD. Based on these studies, a putative model for VD action on cell proliferation and differentiation is presented.

Responses of coxsackievirus B4-specific T-cell lines to 2C protein-characterization of epitopes with special reference to the GAD65 homology region.

Coxsackie B viruses (CBV) have been indicated as environmental triggers initiating autoimmune destruction of insulin-producing pancreatic beta-cells, and molecular mimicry might be the mechanism. A prime candidate for inducing cross-reactive immune responses is a homology sequence, PEVKEK, found both in CBV4 2C protein and in GAD65. To characterize the CBV4-specific T-cell epitopes, overlapping peptides covering the 2C protein were synthesized and CBV4-specific T-cell lines were established from healthy and diabetic subjects. The T-cell epitopes were dependent on the HLA-DR genotype of the T-cell donor, but no difference between diabetic and healthy subjects could be detected. Peptide p4, which included the PEVKEK sequence, contained an HLA-DR1-restricted T-cell epitope. Three randomly selected CBV4-specific T-cell lines, which responded to peptide p4, failed to recognize GAD65 protein or GAD65 peptides containing the PEVKEK sequence. We conclude that the CBV4 2C protein is strongly immunogenic for T-cells and PEVKEK is included in a T-cell epitope. However, presentation of this epitope in the context of neutral HLA-DR1 allele does not support its role in pathogenesis of type 1 diabetes.

Local challenge on oral mucosa with an alpha-gliadin related synthetic peptide in patients with celiac disease.

OBJECTIVE: Gluten-derived peptides (e.g., amino-acids 31-49 of alpha-gliadin) have been shown to cause changes typical of celiac disease in the gut. Gluten-derived peptides have mostly been used in in vitro studies. The easiest access to the gastrointestinal system may be the mouth. In the present study we were interested to see whether a synthetic peptide corresponding to amino-acids 31-49 of alpha-gliadin could induce inflammatory changes in the oral mucosa after a local challenge in celiac disease patients. METHODS: The challenge was made by injecting the peptide solution at a concentration of 10 microg/ml submucosally into the oral mucosa of 10 celiac disease patients after a gluten-free diet (GFD) and 12 healthy control subjects. B and CD45RO+ T cells, mast cells, CD3+, CD4+, CD8+ lymphocytes, and alphabeta and gammadelta T-cell receptor-bearing (TcR alphabeta, TcR gammadelta) lymphocytes were counted and HLA DR expression was determined. The expression of CD25 and Ki-67 antigen was also examined. RESULTS: The peptide significantly increased the total number of T cells in the lamina propria of the celiac disease patients. The expression of T-cell activation marker CD25 (IL-2 receptor), but not that of cell proliferation marker Ki-67, was also significantly increased in the lamina propria after peptide challenge. Such a reaction was not observed in the controls. The numbers of CD3+ and CD4+ T cells in the lamina propria were also increased in celiac disease patients after the challenge. The count of TcR gammadelta+ cells was very small in the oral mucosa in celiac disease and showed no increase when the oral mucosa was challenged with the peptide. The expression of HLA DR staining was enhanced after the submucosal peptide challenge in celiac disease; however, the difference was not statistically significant. CONCLUSIONS: The results show that in the celiac disease patients after the peptide challenge the oral mucosal lamina propria responds with a nonproliferative increase of lymphocytes. Thus, submucosal challenge with the peptide 31-49 can be used as an aid in the diagnosis of celiac disease. However, further studies with optimized methodology, including various concentrations of the peptide, adjuvants, other peptides, etc., are warranted, especially because the oral mucosa provides the easiest access to an in vivo peptide challenge in celiac disease.

The effects of glycopyrrolate on oral mucous host defenses in healthy volunteers.

UNLABELLED: We studied the effects of glycopyrrolate on oral mucous host defenses. Single IV doses of glycopyrrolate (4 microg/kg) or placebo were administered to 12 healthy volunteers in a randomized, double-blinded, cross-over study. Salivary flow rates and the concentrations/activities of total protein, amylase, and nonimmunologic (lysozyme, lactoferrin, myeloperoxidase, total salivary peroxidase, and thiocyanate) and immunologic (total immunoglobulin A, immunoglobulin G, and immunoglobulin M) mucous host defense factors were determined for paraffin-stimulated whole saliva before and 1, 3, 6, 12, 24, and 48 h after drug administration. Glycopyrrolate serum concentrations were determined before and 2, 4, 6, 10, 15, and 30 min and 1, 2, 3, 6, 12, and 24 h after IV drug injection. Salivary flow rates were decreased significantly for 12 h after glycopyrrolate injection, compared with saline injection. The concentrations of immunologic and nonimmunologic defense factors were increased in the glycopyrrolate group, and differences between the groups were found for all factors (P < 0.05-0.001) except lysozyme and total salivary peroxidase. In contrast, because of the reduced flow rate, the output of all defense factors into the saliva was decreased after glycopyrrolate injection, compared with saline injection. Glycopyrrolate thus decreases the output of salivary host defense factors into the oral cavity. IMPLICATIONS: Glycopyrrolate induces long-lasting hyposalivation and decreases the secretion of salivary immunologic and nonimmunologic defense factors in healthy volunteers.

Granulocyte macrophage-colony stimulating factor (GM-CSF) and sucralfate in prevention of radiation-induced mucositis: a prospective randomized study.

PURPOSE: To compare subcutaneously given molgramostim (GM-CSF) and sucralfate mouth washings to sucralfate mouth washings in prevention of radiation-induced mucositis. METHODS AND MATERIALS: Forty head and neck cancer patients were randomly assigned to use either GM-CSF and sucralfate (n = 20) or sucralfate alone (n = 20) during radiotherapy. Sucralfate was used as 1.0 g mouth washing 6 times daily after the first 10 Gy of radiotherapy, and 150-300 microg GM-CSF was given subcutaneously. The grade of radiation mucositis and blood cell counts were monitored weekly. Salivary lactoferrin was measured as a surrogate marker for oral mucositis. RESULTS: We found no significant difference between the molgramostim and the control groups in the oral mucositis grade, oral pain, use of analgesic drugs, weight loss, or survival. The median maximum neutrophil counts (median, 9.2 x 10(9)/L vs. 5.9 x 10(9)/L, p = 0.0005), eosinophil counts (median, 1.3 x 10(9)/L vs. 0.2 x 10(9)/L, p = 0.0004), and salivary lactoferrin concentrations were higher in patients who received GM-CSF. The most common toxicities in the GM-CSF plus sucralfate group were skin reactions at the GM-CSF injection site (65%), fever (30%), bone pain (25%), and nausea (15%), whereas the toxicity of sucralfate given alone was minimal. CONCLUSION: We found no evidence indicating that subcutaneously given GM-CSF reduces the severity of radiation-induced mucositis.

Effects of the common cold and intranasal fluticasone propionate treatment on mucosal host defense assessed by human saliva.

OBJECTIVE: The purpose of this investigation was to study the effect of a potent topical steroid, fluticasone propionate, on patients with early signs and symptoms of the common cold. To characterize the mucosal inflammatory response, salivary defense factors and flow rate in these patients were analyzed. STUDY DESIGN: Forty patients with symptoms of the common cold were randomized into 2 groups to receive either high-dose fluticasone propionate (100 microg per nostril) or placebo 4 times daily for 6 days. Paraffin-stimulated whole saliva was collected on day 1 (before the onset of medication), day 7 (posttreatment), and day 21 (follow-up). RESULTS: Salivary flow rate, innate host defense factors, and total protein content were not affected by the common cold. IgA increased between day 7 and day 21 (P < or = .01; Student 2-tailed t test), and the relative proportions of salivary peroxidase and IgA increased on day 7 (P = .01) and day 21 (P= .05). In patients receiving fluticasone, saliva flow rate was lower on day 21 (P < or = .05) than on days 1 and 7. The innate salivary defense factors were not affected, but IgA increased both on day 7 (P < or = .001) and on day 21 (P < or = .001) in comparison with day 1. CONCLUSIONS: Of the oral mucosal defense factors, only IgA is activated during the common cold. Intranasally administrated fluticasone propionate does not have a suppressive effect on salivary antimicrobial capacity.

T cell epitopes of a lipocalin allergen colocalize with the conserved regions of the molecule.

In this study we characterized the human T cell-reactive sites of the major cow dander allergen, Bos d 2, a member of the lipocalin protein family. We showed that Bos d 2 contains only a limited number of epitopes. This is in contrast to many other allergens, which usually contain multiple T cell epitopes throughout the molecule. The epitopes of Bos d 2 were primarily concentrated in the conserved regions of the molecule. One of the epitopes was recognized by all the cow-asthmatic individuals regardless of their HLA phenotype. Computer-predicted T cell epitopes on Bos d 2, other lipocalin allergens, and human endogenous lipocalins were situated in similar locations on these molecules and corresponded to experimentally identified epitopes on Bos d 2. The results suggest that human endogenous lipocalins could be involved in the modulation of immune responses against exogenous lipocalin allergens. In addition, our findings are likely to facilitate the development of new forms of immunotherapy against allergies induced by the important group of lipocalin allergens.

Association of ezrin with intercellular adhesion molecule-1 and -2 (ICAM-1 and ICAM-2). Regulation by phosphatidylinositol 4, 5-bisphosphate.

Ezrin is a cytoplasmic linker molecule between plasma membrane components and the actin-containing cytoskeleton. We studied whether ezrin is associated with intercellular adhesion molecule (ICAM)-1, -2, and -3. In transfected cells, ICAM-1 and ICAM-2 colocalized with ezrin in microvillar projections, whereas an ICAM-1 construct attached to cell membrane via a glycophosphatidylinositol anchor was uniformly distributed on the cell surface. An interaction of ICAM-2 and ezrin was seen by affinity precipitation, microtiter binding assay, coimmunoprecipitation, and surface plasmon resonance methods. The calculated KD value was 3.3 x 10(-7) M. Phosphatidylinositol 4, 5-bisphosphate (PtdIns(4,5)P2) induced an interaction of ezrin and ICAM-1 and enhanced the interaction of ezrin and ICAM-2, but ICAM-3 did not bind ezrin even in the presence of PtdIns(4,5)P2. PtdIns(4, 5)P2 was shown to bind to cytoplasmic tails of ICAM-1 and ICAM-2, which are the first adhesion proteins demonstrated to interact with PtdIns(4,5)P2. The results indicate an interaction of ezrin with ICAM-1 and ICAM-2 and suggest a regulatory role of phosphoinositide signaling pathways in regulation of ICAM-ezrin interaction.

Stimulated salivary flow rate and composition in asthmatic and non-asthmatic adults.

The number of decayed, missed and filled permanent teeth (DMFT), the degree of periodontal inflammation (Periodontal Status Index, PSI), stimulated salivary flow rate and the concentrations of total protein, lactoferrin, lysozyme, myeloperoxidase, salivary peroxidase, calcium, potassium, sodium and thiocyanate in whole saliva of 26 adult asthma patients were compared with those of 33 non-asthmatic controls. The saliva was also analysed for mutans streptococci, lactobacilli, total anaerobic flora and Candida spp. The mean PSI (p < 0.05; 95% confidence interval for the difference between means (95% CI) 2.47-25.30) was higher and the mean stimulated salivary flow rate (p < or = 0.05; 95% CI 0.57-0.55) was lower in the asthmatic group than in the control group. No differences were found between the groups in non-immune defense factors, except for myeloperoxidase. The myeloperoxidase concentrations were higher in asthmatics than in non-asthmatics (p < 0.05; 95% CI 4.4-134.0 ng/ml). No differences in microbial counts were found. It was concluded that stimulated salivary flow rates decrease while myeloperoxidase concentrations increase in adult asthmatic patients compared with non-asthmatic adults. The higher concentrations of myeloperoxidase are explained by a higher PSI in asthmatics.

Longitudinal analysis of the association of human salivary antimicrobial agents with caries increment and cariogenic micro-organisms: a two-year cohort study.

Previous studies of the possible associations of salivary antimicrobial agents with dental caries have given controversial results, obviously mainly because almost all studies have been cross-sectional. Our aim was to find out, in a two-year longitudinal follow-up study, the associations among selected salivary non-immune and immune antimicrobial variables, cariogenic bacteria, and caries increment. The study population was comprised of 63 subjects, all of whom had their 13th birthday during the first study year. In addition to a comprehensive dental examination at baseline and after 2 yrs, paraffin-stimulated whole saliva samples were collected in a standardized way at six-month intervals. Saliva samples were analyzed for flow rate, buffer effect, lysozyme, lactoferrin, total peroxidase activity, hypothiocyanite, thiocyanate, agglutination rate, and total and specific anti-S. mutans IgA and IgG, as well as for numbers of total and mutans streptococci, lactobacilli, and total anaerobic bacteria. Cluster analysis and Spearman-Rank correlation coefficients were used to explore possible associations between and among the studied variables. During the two-year period, a statistically significant increase was observed in flow rate, thiocyanate, agglutination rate, anti-S. mutans IgA antibodies, lactobacilli, and total anaerobes, whereas lysozyme, lactoferrin, and total and anti-S. mutans IgG antibodies declined significantly. Based on various analyses, it can be concluded that, at baseline, total IgG and hypothiocyanite had an inverse relationship with subsequent two-year caries increment, anti-S. mutans IgG antibodies increased with caries development, and mutans streptococci and lactobacilli correlated positively with both baseline caries and caries increment. Total anaerobic microflora was consistently more abundant among caries-free individuals. In spite of the above associations, we conclude that none of the single antimicrobial agents as such has sufficiently strong power to have diagnostic significance in vivo with respect to future caries.

Immunochemical characterization of a novel mitochondrially located protein encoded by a nuclear gene within the DFNB8/10 critical region on 21q22.3.

A novel protein encoded by the C210RF2 gene in chromosomal locus 21q22.3 was characterized by immunochemistry. This chromosomal region is known to contain genes for human diseases such as non-syndromic autosomal recessive deafness (DFNB8/10) and autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED). Polyclonal murine antisera were produced against the multivalent peptides deduced from the amino acid sequence of the polypeptide. Immunological reactivity of the obtained antisera was tested with primary cells or established cell lines. On western blotting, the polyclonal sera recognized a single protein product of 25 Kd expressed in cell lines of epithelial and lymphoid origin. Subsequent immunochemistry of several human tissues indicated the ubiquitous expression of the protein. Immunofluorescence studies and co-staining with a mitochondrial-specific dye suggest the subcellular localization of the protein to mitochondria. Mitochondrial localization is also predicted by computer analysis of the polypeptide sequence. As deafness is known to be caused in some instances by defects in mitochondrial function, C210RF2 is a plausible candidate gene for DFNB8/10.

Diagnostic phase antibody response to the human papillomavirus type 16 E2 protein is associated with successful treatment of genital HPV lesions with systemic interferon alpha-2b.

BACKGROUND AND OBJECTIVE: Systemic interferon alpha-2b treatment reduces relapses of genital human papillomavirus (HPV) lesions in some but not all females. The aim of the present study was to investigate possible predictive pretreatment factors for the outcome of therapy. MATERIAL AND METHODS: HPV DNA status and HPV antibody response were evaluated in 100 randomized patients treated with laser ablation and systemic interferon alpha-2b or placebo, and followed up to 6 months. RESULTS: Overall, adjuvant therapy with systemic interferon-alpha did not differ from placebo. However, detectable diagnostic phase levels of serum antibodies to e.g. HPV16 open reading frame (ORF) E2 derived peptide 141EEASVTVVEGQVDYY155 predicted 10-fold difference in the risk of recurrence of HPV infection following adjuvant interferon alpha-2b therapy as compared with placebo (odds ratio, OR, 0.5, 95% confidence interval (CI), 0.1-2.3; OR, 4.6, 95% CI 0.5-41, respectively). This trend was statistically significant in the whole study population (2P < 0.05), and in patients with high viral load (2P < 0.01). CONCLUSIONS: Evaluation of the E2 antibody responses may help to identify women with genital HPV lesions who respond to systemic interferon alpha-2b treatment.

Characterization of a nuclear localization signal of canine parvovirus capsid proteins.

We investigated the abilities of synthetic peptides mimicking the potential nuclear localization signal of canine parvovirus (CPV) capsid proteins to translocate a carrier protein to the nucleus following microinjection into the cytoplasm of A72 cells. Possible nuclear localization sequences were chosen for synthesis from CPV capsid protein sequences (VP1, VP2) on the basis of the presence of clustered basic residues, which is a common theme in most of the previously identified targeting peptides. Nuclear targeting activity was found within the N-terminal residues 4-13 (PAKRARRGYK) of the VP1 capsid protein. While replacement of Arg10 with glycine did not affect the activity, replacement of Lys6, Arg7, or Arg9 with glycine abolished it. The targeting activity was found to residue in a cluster of basic residues, Lys5, Arg7, and Arg9. Nuclear import was saturated by excess of unlabelled peptide conjugates (showing that it was a receptor-mediated process). Transport into the nucleus was an energy-dependent and temperature-dependent process actively mediated by the nuclear pores and inhibited by wheat germ agglutinin.

Binding of the cytoplasmic domain of intercellular adhesion molecule-2 (ICAM-2) to alpha-actinin.

Intercellular adhesion molecule-2 (ICAM-2) functions as a ligand for lymphocyte function-associated antigen-1 (LFA-1) and is involved in leukocyte adhesion. We studied intracellular associations of ICAM-2 using a peptide encompassing the cytoplasmic amino acids 231-254 as an affinity matrix. Among the proteins from placental lysates that bound to the peptide was alpha-actinin as demonstrated by immunoblotting. Purified, 125I-labeled alpha-actinin also bound to the peptide. Confocal microscopic analysis of Eahy926 cells demonstrated a colocalization of ICAM-2 and alpha-actinin. Of overlapping octapeptides covering the entire ICAM-2 cytoplasmic amino acids, ICAM-2241-248 bound alpha-actinin most avidly and effectively competed with the longer cytoplasmic peptide for binding. The site of interaction in alpha-actinin was studied using bacterially expressed alpha-actinin fusion proteins. Several constructs covering nonoverlapping regions of alpha-actinin bound to the ICAM-2 cytoplasmic peptide suggesting that multiple regions in alpha-actinin can mediate the interaction. These results, together with previously demonstrated interactions between alpha-actinin and the adhesion proteins ICAM-1, L-selectin, beta1- and beta2-integrins emphasize the role of alpha-actinin as a linker between cell surface adhesion molecules and the actin-containing cytoskeleton.

Autoantibodies against oxidized LDL do not predict atherosclerotic vascular disease in non-insulin-dependent diabetes mellitus.

Accelerated atherosclerosis in diabetes has been suggested as being due to an enhanced oxidative modification of LDL. We hypothesized that the titers of autoantibodies against oxidized LDL (oxLDL) may be increased in patients with non-insulin-dependent diabetes mellitus (NIDDM) and that they may contribute to various manifestations of atherosclerosis among such patients. In a 10-year follow-up study of 91 newly diagnosed NIDDM patients and 82 nondiabetic control subjects, autoantibodies against oxLDL (expressed as the ratio of autoantibodies against oxLDL and native LDL) were measured at baseline and after 10 years. Quantitative ultrasonography to examine the intimal-medial thickness of the common carotid artery (a morphological index of arterial wall injury) and carotid bifurcation was performed at the 10-year examination. The relationship of autoantibodies against oxLDL to the occurrence of cardiovascular death, fatal and nonfatal myocardial infarction, stroke, and any cardiovascular event as well as to the intimal-medial thickness of the common carotid artery and carotid bifurcation was evaluated. Associations between these autoantibodies and metabolic variables (fasting glucose, glycosylated hemoglobin, insulin, and serum lipids) in NIDDM patients were also examined. Autoantibodies against oxLDL did not differ between NIDDM and control subjects (NIDDM: baseline, 1.63 and 0.61 to 23.6; 10-year examination, 1.64 and 0.06 to 59.0; control group: baseline, 1.84 and 0.13 to 36.0; 10-year examination, 1.50 and 0.25 to 8.29; median and range, P = .62, baseline; P = .45, 10 year). In both groups, the titers of these autoantibodies measured at baseline and after 10 years significantly correlated with each other (r = .63 for the diabetic and r = .51 for the control group, respectively, P < .001 for each). The frequency of all cardiovascular events was markedly higher in the NIDDM group than in the control group, but autoantibodies against oxLDL had no significant association with any of these events, including cardiovascular mortality. At the 10-year examination the intimal-medial thickness of the common carotid artery (1.24 +/- 0.36 versus 1.06 +/- 0.30 mm, P = .002) and carotid bifurcation (2.11 +/- 0.73 versus 1.77 +/- 0.82 mm, P = .01) were greater in NIDDM patients than in control subjects, but autoantibodies did not show any association with the intimal-medial thicknesses in either the diabetic or control groups. Autoantibodies against oxLDL indicate the presence of oxidatively modified LDL in vivo, but their titers in the serum do not seem to associate with the excess cardiovascular mortality, morbidity, or intimal-medial thickness of the carotid artery.

Longitudinal analysis of human salivary immunoglobulins, nonimmune antimicrobial agents, and microflora after tonsillectomy.

In order to study the role of tonsils in the host defense in the oral region one pre- and two postoperative (1 and 6 months) whole saliva samples were collected from 25 young adults referred for tonsillectomy. Saliva samples were analyzed for selected host defense factors, representing both immune (total IgA, IgG, IgM, anti-Streptococcus mutans, anti-EBV, anti-CMV, and anti-adenovirus IgA and IgG) and nonimmunoglobulin (lysozyme, lactoferrin, salivary peroxidases, thiocyanate, hypothiocyanite, and agglutinins) mediators. Following tonsillectomy, a significant (P < 0.04) reduction was observed in specific IgG antibodies, suggesting that tonsils participate in local IgG response to oral antigens. Total IgM levels also decreased (P< 0.006), which may to some extent reflect reduced antigenic stimuli compared to preoperative status with frequent tonsillitis. Saliva-derived nonimmunoglobulin host defense factors, except lactoferrin, which declined significantly, remained normal throughout the study period. Our study indicates that tonsils play a role in local oral IgG-mediated immune response but tonsillectomy does not seem to lead to any significant long-term impairment of salivary defense capacity.

Peroxidases, lactoferrin and lysozyme in peripheral blood neutrophils, gingival crevicular fluid and whole saliva of patients with localized juvenile periodontitis.

OBJECTIVE: The aim of this study was to examine the longitudinal association of selected non-immune anti-microbial host factors (peroxidases, lysozyme and lactoferrin) to the localized juvenile periodontitis (LJP) disease status. MATERIALS AND METHODS: Peroxidases, lysozyme and lactoferrin were quantitated from seven patients with LJP before and after periodontal therapy. Analyses were performed from simultaneously collected samples of peripheral blood polymorphonuclear leukocytes (PMNs), gingival crevicular fluid (GCF from diseased sites) and paraffin-stimulated whole saliva. Similar assays were done also from seven periodontally healthy controls. RESULTS: During untreated phase of LJP myeloperoxidase, lysozyme and lactoferrin concentrations were remarkably elevated in peripheral blood PMNs, also reflected in their high concentrations in GCF. All these values normalised with respect to healthy controls during the periodontal therapy. No similar longitudinal changes were seen in whole saliva but during therapy salivary peroxidase concentrations declined below the control values, in accordance with our previous observations in parotid saliva samples of LJP patients. CONCLUSIONS: In LJP the concentrations of lysozyme, lactoferrin and myeloperoxidase are significantly elevated in peripheral blood PMNs, also reflected in GCF. During periodontal therapy these values decline and approach those observed in healthy controls. No similar changes are seen in stimulated whole saliva.

Immunocytochemical evidence for differential subcellular localization of 5 alpha-reductase isoenzymes in human tissues.

To contrast the previously described distribution of 5 alpha-reductase isoenzyme 2 in human organs we have raised synthetic C-terminal peptides of human 5 alpha-reductase isoenzyme 1 (h 5 alpha r1) and isoenzyme 2 (h 5 alpha r2) respectively, and studied the cellular and subcellular distribution of both isoforms of this key enzyme of testosterone metabolism using the human prostate gland as a reference. h 5 alpha r1, which in Western blots of human prostatic proteins had an apparent molecular weight of 23 kDa, was localized immunohistochemically in the nucleus of prostatic epithelial and stromal cells. Ultrastructurally, it was closely associated with the nuclear matrix. The apparent molecular weight of h 5 alpha r2 was 26 kDa in Western blotting of human prostatic proteins. In immunohistochemically processed sections, it was seen in the cytoplasm of prostatic epithelial as well as stromal cells. An organ screening with genital and extragenital tissues of males and females was performed and to elucidate the distribution of the isoenzymes. Presence of both isoenzymes was demonstrated for a number of tissues, including the central nervous system, the urogenital tract, the respiratory tract, the gastrointestinal tract, skin, and the endocrine system. The divergent localization of 5 alpha-reductase isoenzymes points to potentially different functions in various organs. In view of the nuclear localization of isoenzyme 1, its close spatial relationship with the androgen receptor is presumed to indicate a close association with the receptor mechanism.

Newer saliva collection methods and saliva composition: a study of two Salivette kits.

UNLABELLED: Saliva is frequently used as a diagnostic fluid and several collection devices have been developed. OBJECTIVE: The aim of the present study was to investigate the validity and reliability of two types of Salivette collection kits (non-covered cotton roll and polypropylene covered polyether roll) relative to conventional collection of saliva using paraffin wax chewing stimulation. MATERIALS AND METHODS: Whole saliva samples were collected from 16 healthy volunteers. Following a cross-over design saliva was collected in a standardized way. The flow rate was determined and saliva samples were analyzed for pH, buffer capacity, electrolytes and protein/glycoprotein content. RESULTS: We find that Salivette methods do not allow evaluation of flow rate. pH was unaffected but buffer capacity was lower in Salivette collected than in paraffin wax-stimulated saliva. The non-covered cotton rolls reduced the content of Na+, K+, Cl-, as well as glycoprotein markers (hexosamines, fucose, sialic acid), lysozyme, lactoferrin, salivary- and myeloperoxidase but increased the concentrations of Ca2+, PO4(3)- and SCN-. Polypropylene covered polyether rolls affected saliva composition less than the non-covered cotton rolls. Thus, SCN- and sIgA concentrations were higher and lysozyme activity lower in the former (covered roll) saliva than in paraffin wax saliva. The reliability of the Salivette kits was good. CONCLUSION: We conclude that the Salivette method generates data significantly different from conventional paraffin wax-stimulated saliva such as buffer capacity and several electrolytes and organic components. Care should be taken in interpreting the results when such methods are employed.