Biomedical subjects
R Hein
Publications and source records attributed to R Hein.
[MIA ("melanoma inhibitory activity"). Biological functions and clinical relevance in malignant melanoma].
The protein MIA was identified and isolated from the tissue culture supernatant of melanoma cells in vitro by its ability to inhibit thymidine incorporation by melanoma cell lines. After purification and partial sequencing of the peptide, a fragment of the MIA cDNA was cloned by RT-PCR. This cDNA fragment was used to screen phage libraries and subsequently fully encoding human and murine MIA cDNA and genomic DNA clones were obtained. The MIA gene spans a region of approximately 2 kb and is divided into 4 exons. Mapping the MIA gene revealed that the human gene is located on chromosome 19 and the murine gene on chromosome 7. The MIA open reading frame spans 131 (human) or 130 (murine) amino acids. The first 24 (human) or 23 (murine) amino acids represent a signal sequence directing the secretion of MIA into the extracellular compartment. The mature, secreted MIA consists of 107 amino acids and its MW is approximately 11 kDa. Preliminary structural data suggests that MIA is a small globular protein stabilized by two intramolecular disulfide bonds. Expression studies of protein und mRNA levels indicate that MIA is expressed specifically by malignant melanoma cells and chondrocytes. This points to a highly restricted expression pattern which is controlled by the MIA promoter. In addition, MIA provides a clinically useful parameter in patients with malignant melanoma. Enhanced values were measured in the serum of all patients with metastatic melanoma (stage III and IV). In vitro and in vivo experiments using recombinant MIA protein revealed that MIA specifically inhibits attachment of melanoma cells to fibronectin and laminin. Further analysis indicated a direct binding between MIA and the matrix proteins. This finding provides an explanation for the capability of MIA to inhibit proliferation of melanoma cells in vitro. Our studies suggest a putative function of MIA in regulated detachment of melanoma cells from the extracellular matrix which is an important step in metastasis.
In vitro modulation of human melanoma cell invasion and proliferation by all-trans-retinoic acid.
Invasive growth and formation of metastases involve complex interactions between tumour cells, host cells and components of the extracellular matrix. Retinoids, a group of vitamin A derivatives, modulate cell growth and differentiation and have been found to suppress tumour cell invasion in vitro and formation of metastases in vivo. The aim of our study was to investigate changes in proliferation and invasion through membrane barriers in vitro of seven human melanoma cell lines, established from human primary melanomas or metastases, in response to treatment with retinoic acid (RA). These changes were compared with the expression regulation of molecules that have been identified as targets of RA-mediated signal pathways. Invasiveness in vitro was correlated with the origin of the cell lines and was significantly higher in the lines derived from metastases. In all the cell lines proliferation and chemotaxis were inhibited by 10(-5) M RA, but the cell lines established from metastases were significantly more sensitive with respect to inhibition of invasion by RA. The specific expression patterns of MMP-1 and TIMP-2 were detected and regulated by RA in almost all cell lines, whereas expression of MMP-2 and TIMP-1 was not influenced by RA treatment. The most striking difference between the cell lines was a strong downregulation of transforming growth factor-beta (TGF-beta) expression in cell lines derived from metastases when treated with RA in contrast to cell lines from primary melanomas. These data provide evidence that RA modulates growth, chemotaxis and invasion in a broad panel of melanoma cell lines derived both from primary non-metastasized melanomas and metastases. However, distinct molecular mechanisms are involved in mediating these effects.
[Gynecologic operations in fibromyalgia syndrome. A retrospective analysis of 890 patients of a rheumatologic and general practice].
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Melanoma-inhibiting activity, a novel serum marker for progression of malignant melanoma.
Melanoma-inhibiting activity (MIA) was isolated previously as a small soluble protein secreted from malignant melanoma cell lines in vitro. In vivo, highly restricted expression patterns in melanocytic tumors were identified. We therefore quantitated serum levels of MIA protein by means of a nonradioactive ELISA and investigated whether MIA provides a clinically useful parameter in patients with malignant melanomas. Here, we report enhanced MIA serum levels in 13 and 23% of patients with stage I and II disease, respectively, and in 100% with stage III or IV disease. Compared with S-100 and soluble intercellular adhesion molecule 1 serum levels in these patients, MIA was the most sensitive marker. Response to therapy in stage IV disease correlated with changes in MIA serum levels. Measuring repeatedly sera of 350 patients with a history of stage I or II melanoma during follow-up, we detected 32 patients developing positive MIA values. At the time of serum analysis, 15 of them had developed metastases, and one presented with metastatic disease 6 months later. In contrast, none of the patients with normal MIA serum levels developed metastases during the follow-up period of 6-12 months. In conclusion, MIA represents a novel serum marker for systemic malignant melanoma revealing the highest sensitivity and specificity among currently available markers. Useful clinical applications include staging of primary melanomas, detection of progression from localized to metastatic disease during follow-up, and monitoring therapy of advanced melanomas.
Systemic scleroderma. Multicenter trial of 1 year of treatment with recombinant interferon gamma.
OBJECTIVE: To confirm significant improvement of the skin score in systemic sclerosis by treatment with interferon gamma in a larger group of patients and to investigate on a molecular level the influence of interferon gamma on collagen type I messenger RNA expression. DESIGN: Open, noncontrolled multicenter study. SETTING: Five outpatient clinics specializing in the care of systemic scleroderma. PATIENTS: Thirty-two patients suffering from the diffuse or limited form of systemic sclerosis and progressive disease were recruited; 20 patients finished the study. INTERVENTION: Each patient received interferon gamma, 50 micrograms subcutaneously 3 times a week for 1 year. MAIN OUTCOME MEASURE: Skin score, collagen type I messenger RNA in skin biopsy specimens. RESULTS: The patients who completed the study showed an unchanged median skin score after 1 year of therapy. In addition, similar collagen type I messenger RNA levels were detected in skin biopsy specimens taken from involved skin before and after therapy in these patients. CONCLUSIONS: Treatment of systemic scleroderma with interferon gamma is associated with stabilization of the skin score and lack of worsening of visceral involvement.
Anorectal function in systemic sclerosis: correlation with esophageal dysfunction?
PURPOSE: This study was designed to compare esophageal and anorectal function parameters in patients with systemic sclerosis and to define the role of anorectal manometry in the diagnosis of gastrointestinal involvement of systemic sclerosis. PATIENTS AND METHODS: Twenty-six consecutive patients (22 females) with systemic sclerosis originally referred for assessment of esophageal function were evaluated by esophageal and anorectal manometry. Anorectal function parameters were compared between patients with normal and those with disturbed esophageal function. RESULTS: A total of 17 of 26 patients (65 percent) had severe esophageal dysfunction with aperistalsis of the lower two-thirds of the esophagus, whereas 9 patients (35 percent) had normal esophageal manometry. Only three patients (11.5 percent) suffered from occasional fecal incontinence. Anorectal function parameters (resting pressure, maximum squeeze pressure, perception threshold) were not significantly different between patients with normal and those with disturbed esophageal motility. Rectoanal inhibitory reflex was excitable in nearly 90 percent of patients. CONCLUSION: In an unselected group of patients with systemic sclerosis, fecal incontinence and abnormal anorectal function are rather rare findings. Anorectal manometry cannot differentiate between patients with and without gastrointestinal involvement of systemic sclerosis.
Esophageal manometry in systemic sclerosis: screening procedure or confined to symptomatic patients?
The predictive value of esophagus-related symptoms for the diagnosis of esophageal dysmotility induced by systemic sclerosis (SSc) was prospectively evaluated in 50 consecutive patients with SSc. Patients were classified as symptomatic when either dysphagia or repeated episodes of heartburn were present. All patients underwent esophageal manometry; SSc-induced esophageal dysfunction was diagnosed when there was aperistalsis or marked hypocontractility of the distal two-thirds of the esophageal body. Twenty-nine patients (58%) had a history of esophagus-related symptoms, while 21 patients (42%) were asymptomatic. Compared to esophageal manometry, esophagus-related symptoms had a sensitivity of 64%, a specificity of 52%, a negative predictive value of 50% and a positive predictive value of 62% for the diagnosis of SSc-induced esophageal dysfunction. In conclusion, the association of esophagus-related symptoms and esophageal motility pattern is poor. As clinical management strategies depend on proof of esophageal dysfunction, screening examinations are mandatory in all patients with SSc.
Hepatitis B associated liver failure following bone marrow transplantation.
BACKGROUND: Several cases have been reported showing clearance of HBsAg in chronic hepatitis B carriers due to adoptive transfer of immunity by an hepatitis B immunised bone marrow. CASE REPORT: We report on a 27-year-old man with chronic myelocytic leukemia and asymptomatic chronic hepatitis B who received allogeneic bone marrow transplantation (BMT). The donor was his HLA identical brother with natural immunity against hepatitis B. Before BMT the donor had received an additional dose of recombinant hepatitis B vaccine. Twenty days after BMT alanine aminotransferase levels increased and graft versus host disease of the skin was observed. Elevation of liver enzymes was initially attributed to graft versus host disease of the liver and the patient received high doses of steroids in addition to standard immunosuppression. Alanine aminotransferase levels increased up to a maximum on day 52 while the HBV DNA level peaked on day 38 after BMT. A liver biopsy showed reactivation of hepatitis B and treatment with steroids was tapered down. Although alanine aminotransferase and HBV DNA levels decreased, liver function deteriorated. The patient died 130 days after BMT due to liver failure. CONCLUSION: This report indicates that disturbance of the balance between HBV replication and immune control after BMT may result in fatal reactivation of hepatitis B. Careful monitoring, including HBV DNA level and early liver biopsy, of patients with chronic hepatitis B undergoing BMT as well as determination of the HBV immune status of the BMT donor is suggested and necessary.
Double-blind, placebo-controlled study of intralesional interferon gamma for the treatment of localized scleroderma.
BACKGROUND: Localized scleroderma is characterized by circumscribed fibrotic plaques and may progress to widespread skin involvement and fibrosis. Interferon gamma (IFN-gamma) has been shown to be a potent inhibitor of collagen synthesis and of the migration and proliferation of dermal fibroblasts. OBJECTIVE: Our purpose was to determine whether IFN-gamma is effective in the treatment of localized scleroderma. METHODS: A double-blind, randomized, placebo-controlled, multicenter study was conducted. Twenty-four patients with progressive lesions received 100 micrograms of IFN-gamma or placebo subcutaneously on 5 consecutive days for 2 weeks followed by 100 micrograms of IFN-gamma or placebo once weekly for 4 weeks. Thereafter patients were observed for 18 weeks. To determine whether improvement could be related to an altered level of collagen messenger RNA (mRNA), biopsy specimens were taken from uninvolved and involved skin before and after therapy. RESULTS: The patients treated with IFN-gamma or placebo showed no significant difference in size or fibrosis of lesions or collagen type I mRNA synthesis. However, a reduction in the number of new lesions was observed in the IFN-gamma-treated group. The biopsy specimens obtained from involved skin showed a moderate increase of type I collagen and a significant decrease in the small proteoglycan decorin mRNA levels. CONCLUSIONS: The results indicate that IFN-gamma is ineffective in the treatment of localized scleroderma, but may inhibit the development of new lesions.
Acquired perforating collagenosis in a patient with carcinoma of the prostate.
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[Hydroa vacciniforme].
HISTORY AND CLINICAL FINDINGS: For three years, each time the day after intensive exposure to the sun a 5-year-old girl developed moderately painful erythema and blisters on uncovered parts of her body which subsequently healed with scarring. There was no increased photosensitivity among members of her family. There were numerous blisters and bullae, some of them confluent, filled with serous fluid on her back, on her arms an in the face. There were varioliform scars on the extensor surfaces of her arms. INVESTIGATION: There were no porphyrins in red cells, serum or urine. The characteristic skin changes were reproduced on exposure to UV-A. These findings established the diagnosis of hydroa vacciniforme. TREATMENT AND COURSE: Prolonged sun exposure was to be avoided and protective ointment was prescribed together with administration of beta-carotene, 25 mg twice daily. CONCLUSION: In children light-induced skin changes which heal with scarring may be due to hydroa vacciniforme.
Structure and promoter analysis of the gene encoding the human melanoma-inhibiting protein MIA.
We have recently described the isolation of a novel protein, MIA, which is secreted from malignant melanoma cells and elicits growth inhibition on melanoma cells in vitro (Blesch, A., Bosserhoff, A. K., Apfel, R., Behl, C., Hessdörfer, B., Schmitt, A., Jachimczak, P., Lottspeich, F., Schlingensiepen, H., Buettner, R., and Bogdahn, U. (1994) Cancer Res. 54, 5695-5701). Here, we report the structure of the human MIA gene locus, describe its expression pattern in melanocytic tumors in vivo, and provide an initial characterization of the MIA promoter. The MIA gene is encoded by four exons, and the mRNA initiation site was identified 70 base pairs upstream from the translation start codon. MIA mRNA expression in vivo correlated with progressive malignancy of melanocytic lesions and was inducible in other cells by phorbol esters. To investigate mechanisms mediating this melanoma-associated expression pattern, we analyzed the promoter activity of the 1.3-kilobase genomic sequences located 5'-upstream of the MIA gene. The MIA promoter conferred high levels of gene activation specifically in human and murine melanoma cells, and its activity was further enhanced by treatment with phorbol esters. Site-directed mutation of an NF-kB site within the MIA promoter did reduce the basal promoter activity in melanoma cells but did not change significantly enhancement by phorbol esters.
Comparison of migration and invasiveness of epithelial tumor and melanoma cells in vitro.
Tumor invasion of basement membranes is a complex multi-step process. Altered adhesion, as well as increased cell locomotion contribute to tumor cell invasion and metastasis. A variety of in vitro models have been used to measure cell invasiveness. The invasion of basement membranes can be simulated in vitro in blind well Boyden Chambers using the reconstituted basement membrane matrigel or collagen type I as the invasion barrier. The aim of our study was to compare the migration and invasive capacity of epidermal tumor cells (TR 131, TR 146, SCL II, FaDu, HLaC 79) and melanoma cells derived from primary tumors (Mel Ei, Mel Ho, Mel Juso, Mel Wei) or their metastases (Mel Ju, Mel Im, Sk Mel 1, Sk Mel 28). Chemotactic response of epidermal tumor cells was increased toward fibroblast conditioned medium and fibronectin (20 micrograms/ml), while laminin (100 micrograms/ml) stimulated chemotaxis in only 3 epidermal tumor cell lines (HLaC 79, FaDu, TR 146), EGF (10 ng/ml) in only 4 cases (SCL II, FaDu, TR 131, TR 146), and interleukin-1 (IL-1) in only 1 case (FaDu). Epithelial tumor cell conditioned medium had no chemotactic influence on epithelial tumor cells. Fibroblast conditioned medium, fibronectin, EGF and PDGF were potent chemoattractants for all melanoma tumor cells, whereas IL-1 did not induce a significant chemotactic response. While two epithelial tumor cell lines (FaDu, TR 146) were able to penetrate collagen type I, matrigel was an impenetrable barrier for all epithelial tumor cells. Two cell lines from melanoma primary tumors (Mel Ho, Mel Ei) and two cell lines from melanoma metastases (Sk Mel 1, Sk Mel 28) showed no invasion through collagen type I and matrigel, whereas invasion through both barriers could be observed for the metastatic cell lines Mel Ju and Mel Im and in the primary tumor cell line Mel Wei. Therefore, the clinical observation of late and rare metastasis in epithelial tumors and early metastasis in melanoma correlate with our in vitro investigation of invasive behavior in tumor cells. No significant correlation between the invasiveness of melanoma cell lines and their clinical origin could be demonstrated suggesting the existence of subpopulations with varying invasive potential.
Thrombosis of the terminal aorta, deep vein thrombosis, recurrent fetal loss, and antiphospholipid antibodies. Case report.
The antiphospholipid syndrome (APS) consists clinically of both arterial and venous thrombosis, recurrent fetal loss and thrombocytopenia associated with antiphospholipid antibodies (aPL). Most of these patients were initially found to suffer from systemic lupus erythematosus (SLE). There is an increasing group of patients who exhibit antiphospholipid antibodies and thrombotic complications without clinical features of SLE or related autoimmune disease termed primary antiphospholipid syndrome (PAPS). The case of a 29-year-old woman with thrombosis of the terminal aorta and deep vein thrombosis, recurrent fetal loss, antiphospholipid antibodies and serological support for an underlying connective tissue disease, probably preclinical SLE is reported.
Autonomic and sensorimotor neuropathy in patients with systemic lupus erythematosus and systemic sclerosis.
OBJECTIVE: To determine and compare the prevalence and degree of autonomic (ANP) and sensorimotor neuropathy (SNP) in patients with systemic lupus erythematosus (SLE) and systemic sclerosis (SSc). METHODS: Thirty-one patients with SLE and 19 with SSc were investigated. Pupillary ANP was assessed by pupillometry, cardiovascular ANP using a standardized test battery, and SNP by a standardized clinical examination. RESULTS: In patients with SLE (SSc), 26% (10.5%) had a pathological pupillary latency time and 6.5% (13.7%) had an abnormal maximal pupillary area (p for the difference between latency time and maximal pupillary area in SLE = 0.040). Thus, patients with SLE had more often lesions of the pupillary parasympathetic portion of the autonomic nervous system. Overall prevalence of ANP was not different between the 2 groups; 29.0% in SLE and 21.1% in SSc for pupillary ANP and 9.7 and 15.7% for cardiovascular ANP, respectively. Overall prevalence of SNP was 6.5% in SLE and 21.1% in SSc. Disease activity was correlated with ANP (p = 0.008) and SNP (p = 0.020) in SLE. Kidney involvement was associated with more severe ANP in patients with SSc (p = 0.031). Duration of the disease did not correlate with any type of neuropathy. CONCLUSION: ANP and SNP are often present in SLE and SSc. SLE and SSc differ in the pattern of ANP and SNP, which indicates that different structures and neuropathogenic mechanisms may be involved. Patients with SLE had severe pupillary ANP, probably a sign of central nervous system involvement.
A subunit interaction in chloroplast ATP synthase determined by genetic complementation between chloroplast and bacterial ATP synthase genes.
F1F0-ATP synthases utilize protein conformational changes induced by a transmembrane proton gradient to synthesize ATP. The allosteric cooperativity of these multisubunit enzymes presumably requires numerous protein-protein interactions within the enzyme complex. To correlate known in vitro changes in subunit structure with in vivo allosteric interactions, we introduced the beta subunit of spinach chloroplast coupling factor 1 ATP into a bacterial F1 ATP synthase. A cloned atpB gene, encoding the complete chloroplast beta subunit, complemented a chromosomal deletion of the cognate uncD gene in Escherichia coli and was incorporated into a functional hybrid F1 ATP synthase. The cysteine residue at position 63 in chloroplast beta is known to be located at the interface between alpha and beta subunits and to be conformationally coupled, in vitro, to the nucleotide binding site > 40 A away. Enlarging the side chain of chloroplast coupling factor 1 beta residue 63 from Cys to Trp blocked ATP synthesis in vivo without significantly impairing ATPase activity or ADP binding in vitro. The in vivo coupling of nucleotide binding at catalytic sites to transmembrane proton movement may thus involve an interaction, via conformational changes, between the amino-terminal domains of the alpha and beta subunits.
Characterization of selected strongly and weakly invasive sublines of a primary human melanoma cell line and isolation of subtractive cDNA clones.
Invasion of basement membranes is a key step in systemic spread of tumour cells. To analyze genetic mechanisms involved in this process, we have selected strongly and weakly invasive sublines with stable phenotypes from a primary human melanoma cell line by repeated passage through a reconstituted basement membrane in vitro. The sublines differed approximately 5-fold in their invasive potential. Invasiveness correlated with better attachment and overexpression of the integrin alpha v/beta 3 (vitronectin/laminin-receptor). Treatment with retinoic acid inhibited proliferation in both sublines and invasion in the weakly invasive cells but stimulated invasion in the strongly invasive subline. Northern-blot analyses revealed equal levels of mRNA expression regarding collagenase type-IV and retinoic-acid receptors but enhanced expression of TIMP-2 mRNA in weakly invasive cells. The 2 sublines differed significantly in their respective DNA ploidy when compared to the wild-type Mel Im cell line, suggesting that they represent heterogeneous clones present in the primary tumour. We have started to exploit this in vitro system for tumour heterogeneity to clone genes involved in invasion. By a subtractive cDNA cloning strategy, 12 partial cDNA clones were obtained that are specifically overexpressed in the strongly or weakly invasive subline. These results illustrate that stable genetic alterations lead to heterogeneous subpopulations within primary melanomas which differ in their ability to invade basement membranes and interact with components of the extracellular matrix.