S100-Beta, melanoma-inhibiting activity, and lactate dehydrogenase discriminate progressive from nonprogressive American Joint Committee on Cancer stage IV melanoma.

PURPOSE: Monitoring advanced malignant melanoma, serum levels of S100-beta (S100beta) and melanoma-inhibiting activity (MIA) were assessed for the ability to discriminate progressive from nonprogressive disease. S100beta and MIA were supposed to be superior to conventional variables, such as lactate dehydrogenase (LDH) level. PATIENTS AND METHODS: Seventy-one patients with stage IV malignant melanoma according to the criteria of the American Joint Committee on Cancer (AJCC) were included in the study. Results of restaging examinations were used as an independent reference standard for diagnosing progressive disease, and S100beta, MIA, LDH level, and erythrocyte sedimentation rate (ESR) were determined in venous blood just before restaging. Sensitivities and specificities of the parameters were calculated by logistic regression analysis. Discrimination ability was assessed by Somers' D(xy) rank correlation and the area under the receiver-operating characteristic curve (ROC-AUC). RESULTS: All tested serum parameters were significantly elevated in patients with progressive disease. The highest sensitivities according to the established thresholds were found for S100beta and MIA (91% and 88%, respectively). LDH had the highest specificity (92%). ESR was dropped from the analysis because of low specificity. In calculating Somers' D(xy) and ROC-AUC values, S100beta, MIA, and LDH showed high discrimination ability. By multiple logistic regression, LDH was identified to be the only statistically significant marker for progressive disease. S100beta and MIA did not provide additional significant information because of their high correlation with LDH with respect to clinical outcome. CONCLUSION: Elevated serum levels of S100beta, MIA, and LDH indicate current disease progression in AJCC stage IV melanoma. LDH was the most relevant overall parameter.

Subclinical Epstein-Barr virus infection of both the male and female genital tract--indication for sexual transmission.

Epstein-Barr Virus (EBV) can infect B lymphocytes as well as epithelial cells of the oral cavity. Recently, infection of epithelial cells of the inflamed uterine cervix has been demonstrated, and EBV-DNA has been detected in urethral discharge of men suffering from genital infection. We investigated whether EBV can be found in the genital tract of both sexes independently from inflammatory disease states. Genital specimens of men and women of a sexually transmitted diseases outpatient clinic after excluding sexually transmitted diseases and clinically apparent signs of inflammation were investigated using the polymerase chain reaction to screen for EBV-DNA. In 13 of 47 samples (27.7%) swabbed from the uterine cervix, EBV-DNA could be detected. Similarly, 6 of 45 samples (13.3%) scraped from the sulcus coronarius contained EBV-DNA. Our study shows that the female genital tract and likewise the male genital tract can subclinically harbor EBV. These findings suggest i) that in addition to the oral cavity, the female and the male genital tract may be a reservoir for EBV and ii) that sexual transmission of this virus associated with an epidemiology different from that of oral infection may be possible.

Detection of Epstein-Barr virus-DNA in tongue epithelium of human immunodeficiency virus-infected patients.

Oral hairy leukoplakia is a lesion on the lateral part of the tongue that contains replicating Epstein-Barr virus (EBV) and presages progression from human immunodeficiency virus (HIV) infection to AIDS. To clarify the role of EBV in the development of the lesions, we used filter in situ DNA hybridization to determine the prevalence of EBV and of human papillomavirus (HPV) in epithelial cells obtained on swabs from the tongue of HIV-infected patients who had hairy leukoplakia, HIV-infected patients who did not have hairy leukoplakia, and healthy uninfected control persons. In samples collected from the 35 uninfected control persons, EBV DNA could not be detected except at low concentrations in three people. In contrast, all but one of the samples from 11 HIV-infected patients who had hairy leukoplakia contained EBV DNA. Of greatest interest, in 19 of 32 HIV-infected patients who had no signs of hairy leukoplakia, EBV DNA was also detected on the epithelium of the tongue. DNA filter in situ hybridization for the detection of HPV serotypes 6, 11, 16, and 18 in all cases yielded negative results. Statistical analysis showed that the presence of EBV DNA was significantly correlated with the clinical status of the HIV-infected persons, as determined by Walter Reed staging classification, whereas hairy leukoplakia was not. It is concluded that detection of EBV DNA in oral epithelium may be an earlier and more powerful predictor of progression to AIDS than is hairy leukoplakia.

Elevated titers of cell-free interleukin-2 receptor in serum and cerebrospinal fluid specimens of patients with acquired immunodeficiency syndrome.

A sensitive monoclonal antibody based ELISA was used to detect cell-free interleukin-2 receptor (IL-2R) in the body fluids of patients with acquired immune deficiency syndrome (AIDS), a variety of other disease conditions and a control group of apparently healthy (heterosexual and homosexual) males. Two of the 25 control donors showed low titers (1:8) of IL-2 receptor in the serum samples; the cerebrospinal fluid (CSF) specimens from these individuals proved negative. However, serum and CSF specimens from all the 9 patients with AIDS showed significantly elevated titers (range 1:128 to 1:4096) of IL-2 receptor. The presence of moderate titers (range 1:128 to 1:512) of circulating IL-2 receptor could also be detected in all of the 4 patients with acute lymphocytic leukemia. IL-2 receptor was detectable in the CSF and/or serum specimens from 3 of 3 patients with lung cancer, 3 of 4 patients with acute hepatitis B infection, and 2 of 3 patients with multiple sclerosis. IL-2 receptor could not be detected in the serum or CSF specimens originating from patients with legionellosis (3/3), asthma (3/3), or those with non-pulmonary febrile bacterial infections (4/4). It is concluded that soluble IL-2 receptor may be found in serum or CSF specimens from patients with certain (but not all) disease conditions including AIDS. The conspicuously elevated titers of cell-free IL-2R in the body fluids of patients with AIDS may contribute to the drastic impairment of the immune system regulation observed in such patients.

Antigenic domains of the HIV-1 vif protein as recognized by human sera and murine monoclonal antibodies.

To analyze the vif antibody response in individuals infected with the human immunodeficiency virus type 1 (HIV-1) and to determine antigenic epitopes on the vif protein, 104 HIV-1+ sera were screened for reactivity with a recombinant vif protein; 30 (28.8%) of these sera recognized the recombinant vif protein in immunoblot and were employed, together with 17 HIV-1/vif-negative control sera, in an enzyme immunoassay (EIA)-based epitope scanning assay with 183 overlapping decapeptides that covered the complete amino acid sequence of the HIV-1 vif protein (strain BH10). Of the 30 HIV-1/vif+ sera, 87% reacted with decapeptides comprising the two following epitopes: IEWRKKRY (vif amino acids 87-94) or DRWNKPQ (vif amino acids 172-178). The two epitopes were 89% and 100% conserved among different HIV-1 strains and their antigenicity could be confirmed by computer-assisted predictions of vif antigenic determinants. All the sera reactive with recombinant vif protein and with vif peptides originated from patients in CDC stages III or IV. Two murine anti-vif monoclonal antibodies reacted only with the seven C-terminal amino acids of the vif protein (SHTMNGH), which were not recognized by any of the human sera. Our results may be useful for further studies of vif seroreactivity and for the production of anti-vif mono- or polyclonal antibodies using vif peptides.

Cellular immune response to sheep erythrocytes: interrelationship between proliferation of popliteal lymph node cells and footpad swelling.

Mice were sensitized with graded doses of sheep erythrocytes by the intravenous or subcutaneous route and challenged for delayed-type hypersensitivity (DTH) at different times thereafter. The DTH response as assessed by footpad swelling (FPS) was compared to the spontaneous proliferative response of the popliteal lymph node cells (PLNC). Proliferation of PLNC was optimal after sensitization regimens resulting in optimal FPS. The same was true for mice sensitized under cyclophosphamide modulation. Proliferation of PLNC induced by SRBC was antigen-specific, although some crossreactivity with horse red blood cells was observed. Proliferation of PLNC could be abrogated by treatment with anti-Thy-1.2 antiserum plus complement demonstrating the T cell nature of proliferating cells. In accordance with published data, FPS of mice presensitized with a high dose of SRBC as well as FPS of recipients of spleen cells from high-dose-sensitized donors was suppressed. In marked contrast, PLNC proliferation was not diminished in these mice. Although proliferation of PLNC did not parallel FPS under all circumstances, it seems to be a correlate of the cellular immune response to SRBC.

Elevated plasma glutamate levels in colorectal carcinoma patients and in patients with acquired immunodeficiency syndrome (AIDS).

Amino acid concentrations were analyzed in the sera of HIV (LAV/HTLV-III) positive persons and in the plasma of colorectal carcinoma patients. Both groups of persons showed significantly elevated glutamate concentrations when compared with healthy control persons. Glutamate concentrations were found to be strongly elevated in all groups of HIV-positive persons including patients with acquired immunodeficiency syndrome (AIDS) or lymphadenopathy as well as HIV-positive persons without overt symptoms, indicating that increased plasma glutamate levels may be among the earliest consequences of the HIV infection. Moreover, the increased plasma glutamate concentrations in the colorectal carcinoma patients were correlated with a decreased immunological reactivity (mitogenic responses against concanavalin A). This suggests the possibility that the increased plasma glutamate concentrations may be causally responsible for the decreased immunological reactivity in colorectal carcinoma patients as well as in patients with AIDS.

Occurrence of antibodies against chlamydial lipopolysaccharide in human sera as measured by ELISA using an artificial glycoconjugate antigen.

An artificial glycoconjugate containing, as a ligand, the deacylated carbohydrate backbone of a recombinant Chlamydia-specific lipopolysaccharide was used as a solid-phase antigen in ELISA to measure antibodies against chlamydial LPS. The specificity and reproducibility of the assay was shown by using a panel of prototype monoclonal antibodies representing the spectrum of antibodies also occurring in patient sera. These mAbs recognized Chlamydia-specific epitopes [alpha 2-->8-linked disaccharide of 3-deoxy-D-manno-octulosonic acid (Kdo) or the trisaccharide alpha Kdo-(2-->8)-alpha Kdo-(2-->4)-alpha Kdo] or those shared between chlamydial and Re-type LPS (alpha Kdo, alpha 2-->4-linked Kdo disaccharide). The assay was used to measure IgG, IgA and IgM antibodies against chlamydial LPS in patients with genital or respiratory tract infections. In comparison to the results obtained with sera from blood donors, it became evident that both types of infection result in significant changes in the profile of LPS antibodies.

Chlamydial infection--a female and/or male infertility factor?

After screening a large series (n = 491) of asymptomatic males of infertile partnerships for chlamydial immunoglobulin (Ig) G antibodies (Chlam AB), no significant influence of past chlamydial infection was found with regard to semen analysis, postcoital testing, in vitro sperm-cervical mucus penetration tests with hormonally standardized cervical mucus, circulating antisperm antibodies (detected with three different methods), local IgG and IgA antibodies (detected by means of the mixed antiglobulin reaction test) on the sperm surface, the sperm-cervical mucus contact test, and a microbial screening of semen samples for mycoplasmas and other potentially pathogenic micro-organisms. However, when the findings were correlated with infertility factors of patients' female partners and the subsequent pregnancy rate in a prospective study, a significant positive correlation of male Chlam AB with a tubal factor in their wives as cause of the couple's infertility was found. The results suggest that the main influence of Chlamydia trachomatis on male fertility is based on sexual transmission and negative influence on tubal function of female partners, but not on reduced sperm functional capacity.

Alpha 2-macroglobulin and alpha 2-macroglobulin/proteinase complexes in human seminal fluid.

OBJECTIVE: To determine the broad-spectrum proteinase inhibitor alpha 2-macroglobulin (alpha 2M) and its functional subforms, i.e., free alpha 2M and alpha 2M/proteinase complexes, in human seminal fluid by using specific enzyme immunoassay systems. SETTING: The study has been performed in the Andrology Department of the Dermatology Clinics and the Laboratory of Immunopathology of the Institute of Immunology. PATIENTS, PARTICIPATES: Routine patients attending the Andrology Department. INTERVENTIONS: The data have been obtained without particular interventions before or after collection of seminal fluid. MAIN OUTCOME MEASURES: The aim of the study was to determine whether alpha 2M or alpha 2M/proteinase complexes are present in human seminal fluid. RESULTS: The concentration of total alpha 2M in human seminal fluid ranged from 1 to greater than 1,000 ng/mL, and between 56% and 85% of the inhibitor was complexed with proteinases. CONCLUSIONS: These findings show that alpha 2M and alpha 2M/proteinase complexes have to be considered as functionally relevant biomolecules in male genital tract secretions.

Prognosis of metastatic melanoma: no correlation of tyrosinase mRNA in bone marrow and survival time.

Recent publications suggest that tyrosinase mRNA in blood as well as in bone marrow is detectable only in a subgroup of patients with metastatic melanoma. This would imply that tyrosinase mRNA is of limited value as a tumor marker. We addressed the question of whether patients with metastatic melanoma and RT-PCR-detectable tyrosinase mRNA in blood or bone marrow have a different prognosis than tyrosinase mRNA-negative patients. Twenty melanoma patients with widespread clinical metastases were enrolled; the survival time after first diagnosis of visceral metastases was correlated to tyrosinase mRNA presence in blood and bone marrow samples. The time of survival of eight patients with metastatic melanoma and detectable tyrosinase mRNA in either blood or bone marrow was not different from the prognosis of 12 patients without detectable tyrosinase mRNA in either blood or bone marrow. Detection of tyrosinase mRNA in blood or bone marrow samples of melanoma patients with advanced disease seems to have no substantial relevance for survival time and outcome of disease. In this constellation, detection of tyrosinase mRNA by RT-PCR is not a valid tumor marker. Nevertheless, tyrosinase positivity in bone marrow in earlier tumor stages might indicate increased risk for the development of distant metastases. This should be addressed in further studies.

The protein phosphatase 2A subunit Bgamma gene is identified to be differentially expressed in malignant melanomas by subtractive suppression hybridization.

Several genes implicated in the development of various malignancies appear to be of minor relevance in melanoma. We therefore aimed to find a tumour suppressor candidate involved in this malignancy by comparing gene expression in uncultured primary melanoma specimens with those in acquired melanocytic naevi, from which quite often melanomas are known to arise. Applying the subtractive suppression hybridization technique, we generated a subtracted library of candidate genes downregulated in melanoma. Among the cDNA fragments identical to known genes, this library included a cDNA fragment 630 bp in length that is identical to the gene for the human protein phosphatase 2A (PP2A) regulatory subunit B (B56) gamma isoform (PP2A-Bgamma, PPP2R5C). On further evaluation of 15 primary melanoma and 16 acquired melanocytic naevus tissue specimens from independent patients using semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis, expression of this gene was found to be suppressed in melanomas compared with naevi; the difference was statistically significant. As PP2A is known to be a major cellular serine-threonine phosphatase, and has been implicated not only in the regulation of cell growth and division but also in the control of gene transcription and growth factor signal transduction, alterations in the pattern of the regulatory subunits may affect substrate specificity and subcellular localization of the PP2A holoenzyme in melanoma cells.

Are responses to therapy of metastasized malignant melanoma reflected by decreasing serum values of S100beta or melanoma inhibitory activity (MIA)?

In metastatic melanoma S100beta as well as melanoma inhibitory activity (MIA) are elevated in the serum in the majority of patients. Elevation has been found to correlate with shorter survival, and changes in these parameters in the serum during therapy were recently reported to predict therapeutic outcome in advanced disease. However, the value of these markers with respect to other possible markers by multivariate analysis has not yet been proven for individual patients. In this prospective study, S100beta and MIA were measured in the serum of 67 consecutive patients before and following treatment. Analysing both the sensitivity and the specificity of the serum parameters by the areas under the receiver operating characteristics (ROC) curves, decreases in S100beta and MIA during therapy were associated with response to therapy, while increases indicated progressive disease. Unexpectedly, the individual diagnostic value of changes in tumour markers during therapy was not superior to one-point measurements at restaging. Moreover, S100beta and MIA were not superior to the conventional parameters lactate dehydrogenase and C-reactive protein (CRP) on multiple logistic regression analysis. Applying classification and regression trees (CARTs), one-point measurements of CRP was shown to be the most relevant overall parameter.

Detection of Epstein-Barr virus DNA in anal scrapings from HIV-positive homosexual men.

Epstein-Barr virus (EBV) can infect epithelial cells as well as B lymphocytes. Infection of the male and female genital tracts has recently been demonstrated, and it has been suggested that the virus may be sexually transmissible. In our study we investigated whether EBV can be found in the anal region of sexually active homosexual men. Anal scrapings from HIV-positive homosexual men and a heterosexual control population were investigated using the polymerase chain reaction (PCR) to screen for EBV DNA. EBV DNA was detected in 8 of 27 anal samples (29.6%) from the homosexual men and 3 of 34 samples (8.8%) from the heterosexual men. Our study shows that, like the genital tract, the anal region can harbour EBV subclinically. This finding suggests that the anal region may be a reservoir for EBV and that sexual transmission of this virus may be possible.

Plasminogen activation in lesional skin of Pemphigus vulgaris type Neumann.

Zymographic and immunological studies revealed that primarily tissue-type plasminogen activator and to a lesser extent urokinase-type plasminogen activator were present in fluids of pemphigus vulgaris (type Neumann) skin blisters. Furthermore, plasmin activity was detected in pemphigus blister fluids using chromogenic peptide substrate assays. In pemphigus, but not in control, suction blister fluids plasmin/alpha 2-antiplasmin and plasmin/alpha 2-macroglobulin complexes were found by immunoprecipitation or by testing in immunoassays after fractionation by molecular-sieve chromatography. Plasmin activity, detected by a low molecular weight chromogenic peptide assay, was ascribed to plasmin/alpha 2-macroglobulin complexes. Since formation of plasmin/inhibitor complexes requires active plasmin, the finding indicates previous activation of plasminogen in pemphigus lesions.

Combined mechanical and antibiotic periodontal therapy in a case of Papillon-Lefèvre syndrome.

BACKGROUND: Papillon Lefèvre syndrome (PLS) is a rare entity and, as such, it is almost impossible to evaluate an effective therapy in a randomized controlled study. The amount of success reported after therapy for prepubertal periodontitis (PP) in PLS is highly variable from case to case. The goal of this case report is to evaluate the effects of a combined mechanical and antibiotic periodontal therapy regimen in the management of PLS. METHODS: A male patient was diagnosed as suffering from PP associated with PLS at the age of 7 years. He showed hyperkeratosis of the palms and soles, as well as advanced periodontal disease already affecting permanent teeth with maximal probing depth and vertical attachment loss of 12 mm and 11 mm, respectively. Subgingival debridement was performed with simultaneous administration of oral 250 mg amoxicillin 3 times daily and 250 mg metronidazole twice daily for one week. Clinical parameters were assessed and subgingival plaque was collected from all teeth prior to therapy and 7 and 26 months after treatment. Selective cultures for A. actinomycetemcomitans were incubated for each individual tooth and DNA probe analysis was performed for various periodontal pathogens. RESULTS: Prior to combined mechanical and antibiotic treatment, all teeth but one harbored Actinobacillus actinomycetemcomitans subgingivally. However, at 7 and 26 months after therapy A. actinomycetemcomitans could be detected neither by culture nor by DNA probes. Clinical parameters improved markedly and teeth erupting after therapy did not exhibit attachment loss of more than 1.5 mm during the observation period. CONCLUSIONS: Eradication (suppression beneath detection levels) of A. actinomycetemcomitans seems to play a significant role in the successful treatment of localized prepubertal periodontitis in PLS.

Massive lethal cerebral bleeding in a patient with melanoma without intracranial metastasis.

The case history is reported of a patient with melanoma and advanced metastases, who died from massive cerebral bleeding. The lethal event was not caused by intracerebral metastasis but by thrombocytopenia. Depression of the bone marrow resulted from tumour infiltration of the skeleton, chemotherapy and vertebral irradiation. An increase of intracranial pressure triggered the cerebral bleeding, caused by haematemesis from a gastric metastasis directly preceding sudden somnolence.

Seropositivity for MIA and S100 in patients with gastrointestinal carcinomas.

Serum levels of melanoma inhibiting activity (MIA) and S100, both markers in malignant melanoma, are increased only in few patients with non-melanocytic tumors. We examined a series of serum samples from patients with colorectal (CRC) (N=56), gastric (GC) (N=43), pancreatic (PC) (N=29), hepatocellular (HCC) (N=30), cholangiocellular and gallbladder carcinoma (CCC) (N=18). MIA and S100 were measured by commercially available assays. Positive serum levels for MIA and S100 were found in 16.1% and 5.4% of the patients with CRC, 11.6% and 9.3% with GC, 34.5% and 13.8% with PC, 0% and 30% with HCC and 16.7% with CCC, respectively. All patients with sera positive for either MIA or S100 suffered from advanced tumors and received palliative treatment. Elevated serum levels of MIA and S100 are frequent in patients with gastrointestinal cancer. Further investigation is warranted to define the role of MIA or S100 seropositivity in gastrointestinal cancer with regard to follow-up.

Interleukin-6 and its surrogate C-reactive protein are useful serum markers for monitoring metastasized malignant melanoma.

Malignant melanoma cells are known to secrete interleukin-6, and elevated interleukin-6 serum levels were reported to correlate with shorter median survival rates. We, therefore, investigated serum values of interleukin-6 and its surrogate C-reactive protein for the ability to discriminate progressive from non-progressive metastatic melanoma disease. Just prior to re-staging examinations, interleukin-6, C-reactive protein and the conventional parameter lactate dehydrogenase were determined in 74 patients with stage IV malignant melanoma according to the criteria of the American Joint Committee on Cancer. We found all tested serum parameters to be significantly elevated in progressive disease. Calculating sensitivities and specificities by logistic regression analysis, the highest sensitivities, according to the established thresholds, were found for interleukin-6 and C-reactive protein with 86% and 76%, respectively. Lactate dehydrogenase had the highest specificity with 94%. Calculating Somers' D rank correlation and the area under the "Receiver Operating Characteristic" curve, all three parameters showed high ability to driscriminate progressive from non-progressive disease. By multiple logistic regression, lactate dehydrogenase was identified to be the most statistically significant marker for progressive disease. We conclude that, comparable to lactate dehydrogenase, interleukin-6 and its surrogate C-reactive protein are useful serum markers for monitoring metastatic malignant melanoma.

[Current status of vaccine development in sexually transmissible diseases]. / Der gegenwärtige Stand der Vakzineentwicklung bei sexuell übertragbaren Erkrankungen.

A prophylactic vaccine represents a major hope for the control of sexually transmitted diseases. The current general vaccine strategies and the status of vaccine development against infections with Neisseria gonorrhoeae, Treponema pallidum, Chlamydia trachomatis and Herpes simplex virus are described. Vaccines consisting of whole infectious agents are replaced by protective subunits. A subunit vaccine has the advantage to be free from other components, which are not relevant for protection and which may confer unwanted side effects. At the present time vaccine development against infections with Neisseria gonorrhoeae and Herpes simplex virus seems to be the most progressed. With monoclonal antibodies several surface components could be identified, which are of importance for the pathomechanism. With Treponema pallidum and Chlamydia trachomatis the development is delayed by unsolved problems of immunity. For the production of vaccines molecular-biologic methods, like protein synthesis or gene-cloning will be used. Genetically modified live vaccines or polytope hybrid vaccines will gain importance in the future.