Detection of high-risk HPV (16, 18, 33) in situ cancer of the cervix by PCR technique.

OBJECTIVE: The purpose of this study was to collect data about the incidence of high-risk HPV (16, 18, 33) types in in situ cervical cancers, and to evaluate the reliability of the morphological signs of HPV infection by comparing the presence of these signs to the PCR-proven HPV virus infection. METHODS: Fifty patients who underwent conisation at the Department of Obstetrics and Gynecology of Semmelweis University, Budapest, Hungary because of in situ cervical cancer were examined retrospectively for the presence of HPV infection by the PCR technique. The direct and indirect morphological signs of HPV infection identified in the histological and cytological samples were compared to the actual results of virus DNA amplification by PCR in the identical histological sections. The evaluation of the cytological smears and the histological sections was accomplished independently by two different pathologists. RESULTS: E6 open reading frame of HPV 16, 18 or 33 was detected by PCR in 56% (28 cases) of the histological sections of the 50 examined patients with in situ cancer. In 92% (26 patients) of the 28 HPV positive patients one HPV type was detected, while in one of the remaining two cases two HPV types (16/33), or all three types could be detected. The direct morphological signs for HPV infection proved to be 75% sensitive and 50% specific when compared to the results of PCR. Their predictive value for HPV infection was 65%. For the indirect HPV signs the sensitivity was 64% and specificity 31%. The predictive value, prognosticating the presence of HPV 16, 18, 33 infection was 54% in the same sections. Using significance analysis no significant relationship (p = 0.7728) could be detected between the positivity of indirect signs and the presence of HPV 16, 18, 33 infection, while in case of direct signs the relationship was almost significant (p = 0.0675). The joint testing of the direct and indirect signs did not improve the results (p = 0.1338). During the review of the cytological smears the specificity of the cytology in predicting true HPV infections was found to be 68% and sensitivity was 20%. The predictive value was only 50%. A significance analysis was not accomplished by this diagnostic method because of the missing data (see text). CONCLUSION: The method of Nawa et al. seems to be a reliable approach for the detection of HPV DNA in paraffin-embedded material. The three main types of HPV (16, 18, 33) are probably represented in lower percentages in CIN III in Hungary, but a larger survey is needed to obtain reliable data. The direct and indirect morphological signs of HPV infection failed to show a significant relationship with the PCR proven presence of HPV 16, 18, 33.

Expression of decorin, transforming growth factor-beta 1, tissue inhibitor metalloproteinase 1 and 2, and type IV collagenases in chronic hepatitis.

Decorin is a small extracellular matrix proteoglycan. It binds and modulates transforming growth factor (TGF)-beta 1 action, the major stimulator of fibrogenesis. Its role in the pathogenesis of human liver cirrhosis is unknown. Therefore, we studied the relationship of the 2 proteins in normal human liver and in 43 chronic hepatitis and liver cirrhosis specimens. To understand the mechanism that maintains matrix deposition in stage IV hepatitis, we studied expression of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2, as well as the activities of type IV collagenases. Gene expression was analyzed on messenger RNA and protein level by morphologic and biochemical approaches. Decorin proved to be an early marker of fibrogenesis, and its deposition increased parallel to that of TGF-beta 1 and to inflammatory activity. Liver fibrosis progressed despite high temporospatial expression of decorin with TGF-beta 1. Neither decorin nor TGF-beta 1 protein deposition increased further in cirrhosis with low inflammatory activity, suggesting that impaired extracellular matrix catabolism rather than active production plays a role in this stage. This possibility was supported by high message levels of metalloproteinase inhibitors, no 72-kd collagenase activities, and low 92-kd collagenase activities.

Expression of a decorin-like moleculein human melanoma.

Decorin, a member of the family of small leucin-rich proteoglycans, has originally been described as a secreted proteoglycan component of the connective tissues, and has been implicated in the negative regulation of cell proliferation directly or via interactions with TGF-beta. It was reported to be generally absent from tumor cells. Here we show that human melanoma cell lines express a decorin-like molecule. We detected decorin mRNA by RT-PCR in 7 out 7 human melanoma lines characterized by various metastatic potential. Using polyclonal antiserum against the core protein of decorin, the typical 80-120 kD glycanated form as well as a high molecular weight aberrant version (200-210 kD) of decorin were demonstrated by Western blot technique in the culture supernatants as well as in lysates of human melanoma cells. Finally, decorin epitope was also demonstrated immunohistochemically in human melanoma xenografts, as well as in tumor cells of surgically resected melanomas but not in melanocytes of nevi. The expression of this aberrant decorin did not inhibit the in vitroor in vivogrowth of human melanoma cells, and it was independent of their metastatic potential. Human melanoma cell lines expressing aberrant decorin retained sensitivity to the antiproliferative and gelatinase-stimulatory effects of exogenous TGF-beta.

Cytokine regulation of syndecan expression in cells of liver origin.

Syndecan-1 and syndecan-2-two cell surface heparan sulfate proteoglycans-were described in normal human liver. Proteoglycans can modulate the effect of cytokines, and cytokines can influence the expression of proteoglycans. In the present work the regulatory effect of IL-1beta, IL-6, TNF-alpha, IFN-gamma and TGF-beta1 on syndecan-1 and syndecan-2 expression of hepatocytes, hepatoma cell lines, liver and skin fibroblasts has been studied. All cytokines were able to influence the steady state level of syndecan-1 and syndecan-2 mRNA. Their action was target cell specific resulting in either up- or downregulation except TGF-beta1 that was stimulatory in all cell types examined.

Decorin and actin expression and distribution in patients with chronic hepatitis C following interferon-alfa-2b treatment.

BACKGROUND/AIMS: Chronic hepatitis C can lead to cirrhosis and hepatocellular carcinoma. Interferon-alfa therapy may prevent the progression of the disease. The expressions of decorin and alfa-smooth muscle cell actin of the extracellular matrix play a central role in liver fibrosis. We set out to assess the expressions of these proteins in chronic hepatitis C patients, and to evaluate how they can be modified by interferon-alfa therapy. METHODS: Twenty chronic hepatitis C patients received interferon-alfa-2b therapy for 6 months (group I) or 12 months (group II). Liver biopsy samples were taken before and after the therapy. The alfa-smooth muscle actin-positive cells were determined with a monoclonal antibody, and decorin expression was detected with a polyclonal antibody. The cells were evaluated with a semiquantitative scoring method. For statistical analysis, non-parametric methods were used. RESULTS: Before the therapy, alfa-smooth muscle actin-labeled cells and marked decorin expression were present throughout all the acinar zones. Interferon-alfa-2b therapy resulted in significant decreases in both the number of alfa-smooth muscle actin-positive cells and the decorin expression. The alfa-smooth muscle actin-positive cells and decorin expression correlated with the histological activity index (R=0.72, p<0.03, R=0.68, p<0.05). CONCLUSIONS: This study demonstrates that a large number of alfa-smooth muscle actin-positive cells and a marked decorin expression are frequent findings in chronic hepatitis C. Treatment with interferon-alfa-2b for 12 months reduced the number of labeled cells and the decorin expression. The results suggest that interferon-alfa-2b is capable of interfering with fibrogenesis in an early and presumably still reversible phase of chronic hepatitis C.

Expression of syndecan-1 in human B cell chronic lymphocytic leukaemia.

Syndecan-1 is considered an important transmembrane proteoglycan in cell-microenvironment interactions, but its exact function in normal or in transformed B cells is still unknown. In this study, RNA was isolated from peripheral cells of chronic lymphocytic leukaemia (B-CLL) and 'normal', non-leukaemic patients, as controls. Reverse PCR showed no or very low syndecan-1 mRNA expression in controls, while in 11/13 B-CLL the circulating leukaemic cells expressed syndecan-1. Similar results were obtained for interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6). Furthermore, syndecan-1 protein was detected in the majority of circulating B-CLL cells by flow cytometry and immunocytochemistry using anti-syndecan-1 MAb. Control cells were practically negative. Further study is required to understand the biological significance of syndecan-1 on B-CLL cells.

Syndecan-1 - A new piece in B-cell puzzle.

Syndecans are transmembrane proteoglycans, with core proteins mainly decorated with heparan sulfate chains. Syndecan-1 is expressed in a tissue-, cell-and differentiation-specific manner. Its extra-cellular domain can bind via HS chains to matrix elements, to growth factors (especially "heparin-binding" proteins) and to certain biological agents. The ectodomain released by proteolysis can also be functionally active. The cytoplasmic domain can take part in signaling processes as well as in modifying cell shape. In hematopoietic cells syndecan-1 is expressed in normal pre-B-cells and plasma cells, as well as in plasmocytoid and lymphoplasmocytoid malignancies. According to our study syndecan-1 is expressed in B-CLL cells both in tissue environment and in circulation.

Proteoglycan gene expression in rat liver after partial hepatectomy.

Liver regeneration is regulated by several factors. Among these extracellular matrix plays a crucial role in the restoration of normal structure. Proteoglycans are major components of extracellular matrix and they are able to bind growth factors implicated in liver growth. We studied syndecan, perlecan, fibroglycan, and decorin expression after partial hepatectomy in rats by Northern blot analysis. A strong early upregulation was observed at 30 min in decorin expression which was followed by the first syndecan and perlecan peaks at 2 and 4 hours after hepatectomy. At 24 hours, which is the peak of hepatocyte DNA synthesis, all these three proteoglycans had elevated steady state transcript level. Fibroglycan message decreased after partial hepatectomy and remained low during the experimental period. The different expression pattern of the proteoglycans suggests that these extracellular matrix components may selectively influence the regeneration process.

Expression of extracellular matrix proteoglycans perlecan and decorin in carbon-tetrachloride-injured rat liver and in isolated liver cells.

Proteoglycans are important components of the extracellular matrix. They are involved in liver regeneration as well as in liver fibrosis. The distribution and cellular source of proteoglycans under normal as well as pathological conditions is still under debate. Localization of decorin and perlecan was studied in normal, acutely damaged, and cirrhotic liver by histochemistry. Furthermore, their synthesis was analyzed in different liver cell populations isolated from normal rat liver. In normal liver, decorin positivity was observed in the perisinusoidal space and in the portal area. Perlecan was clearly detectable in the portal area (blood vessels and bile ducts); a moderate reaction was also seen along the sinusoids. Strong positivity for both proteoglycans was detectable in the necrotic areas of acutely damaged liver. Chronic liver damage was characterized by the deposition of decorin and perlecan in the fibrotic septa. Immunocytochemical reactions were positive for perlecan and decorin in cultured Ito and endothelial cells but not in hepatocytes and Kupffer cells. Northern hybridization confirmed the capacity of Ito cells and endothelial cells to express the two genes. Interestingly, although rat skin fibroblasts expressed strong messages for both proteoglycans, rat aortic smooth muscle cells did not synthesize decorin.

Syndecan-1 gene expression in isolated rat liver cells (hepatocytes, Kupffer cells, endothelial and Ito cells).

Normal liver contains mainly heparan sulfate proteoglycans. To determine which liver cells are able to express syndecan-1 RNA, hepatocytes, endothelial, Kupffer and Ito cells were isolated from normal rat liver and kept in culture. Rat fibroblasts, monocytes and peritoneal macrophages were also studied. Immediately after isolation the steady state level of syndecan messages in hepatocytes was comparable to that of the normal liver. The expression increased at day one and then remained constant until day five. Ito and endothelial cells contained a low amount of syndecan message. Freshly isolated Kupffer cells failed to express syndecan, but strong upregulation was found on the first day in culture which gradually decreased by day 5. Syndecan transcripts were dose and time dependently upregulated by endotoxin and gamma interferon in Kupffer cells. Endotoxin had no effect on fibroblasts.

[Stiff man syndrome]. / Stiff-man-Syndrom.

Our 51-year old patient developed low back pain and an increase of tonus mainly in the left limb and trunk muscles. Microscopic examination of the muscles and electronmicroscopic examination of the sural nerve and that of the gastronemic muscle did not show any specific deviation. On the basis of spontaneous activity observed during EMG examination, protracted motor activity of great amplitude when moved passively, as well as characteristic clinical symptoms and the disease process Stiff-man syndrome was diagnosed. Considering literature data, we tried applying diazepam (Seduxen and Rivotril)-presumably increasing the praesynaptic inhibition and affecting the reticular system of the brain stem-, as well as GABA medicaments (Lioresal and Depakine) increasing synaptic transport. By giving Seduxen, Rivotril and Baclofen simultaneously a lasting remission of symptoms could be reached. Applying Depakine in combination with the above medicines proved ineffective, presumably because of synergetic side effects.

X-ray diagnostic sign for the differentiation of neurogenic and primary muscular diseases.

The authors give an account of x-ray examinations of the limb musculature of 70 patients suffering from neurogenic muscular diseases, 42 suffering from primary muscular diseases and 45 suffering from senile degeneration of the muscles. Different degree of damage to different parts of the same muscle could only been observed in one case of neurogenic atrophy (in the post-poliomyelitic state) and in two cases of senile degeneration, while it was found in 11 cases (20%) for the other muscular diseases. In the latter cases the more severe muscle damage, which could be demonstrated radiographically, was always found in the part of the muscle adjacent to a tendon. On the above reasons the authors consider that radiographically demonstrable partial or uneven damage to any particular muscle can be used as a new diagnostical information in distinguishing muscular diseases from neurogenic muscular atrophy.

Use of X-ray techniques to demonstrate electively increased damage to certain muscles in patients suffering from muscular diseases.

In X-ray examinations on 38 patients suffering from muscular diseases the authors demonstrated electively pronounced damage to certain muscles in 18 cases. The decay of muscle fibres within a single muscle is not always uniform and is often bilaterally asymmetrical. Before biopsy or EMG examinations it is advisable to take an X-ray of the damaged musculature.

Roentgenmorphological aspects of muscular pseudohypertrophy.

Using special soft tissue X-ray investigations the authors have found that the increase in volume of the calf musculature in progressive muscular dystrophy does not seem to be in complete accordance with the criteria of pseudohypertrophy. It has been demonstrated that this structural alteration of the musculature does not damage the various muscles to an equal extent. It has been shown that in two cases of motoneurone lesions the total decay of the calf musculature was not followed by loss of muscle bulk. The volume of the damaged musculature remained unchanged, or increased, in consequence of the enormous increase in the fat and connective tissue. This structural alteration corresponds to the generally accepted definition of pseudohypertrophy.

Myopathy with hyperaldosteronism. An electron-microscopic study.

In the case reported, an overproduction of aldosterone was accompanied by paretic attacks, a decrease in the serum potassium level, and in muscle tone and the deep tendon reflexes. The decrease of serum potassium level was consistent, but moderate, being to just below the normal lower limit. Loading with carbohydrate produced an attack, forearm ischaemic exercise resulted in less lactic acid production than normally. In the limb-girdle muscles there was also evidence of slight but permanent weakness. The results of histological and ultrastructural examination of a muscle biopsy are reported. It seems probable that the disturbance in the mineralo-corticoid metabolism led periodically to a decrease in the serum potassium, while in some muscle fibres the glycogen content increased and the SR became dilated and partially destroyed. In the attack-free periods tubular aggregates were produced, supposedly through an attempt at regeneration of the SR. During attacks portions of the muscle fibres were destroyed irreversibly and in consequence moderate permanent weakness resulted. Some spheromembranous degradation products which were observed might be regarded as remnants of degenerated tubular aggregates.

Fine structural alterations of muscle fibers in diseases accompanied by myotonia.

The authors have reported the results of examination by electron microscopy of two muscle biopsy specimens from cases of myotonia congenita and three cases of myotonia dystrophica. They have stated that "peripheral annular formation" was a frequently observed alteration in the myotonia congenita cases. In the myotonia dystrophica cases there were additionally disorganized myofibrils in the subsarcolemmal region and inclusion body vacuoles morphologically connected with the sarcolemma. The term "peripheral annular formation" refers to the situation in which the peripheral myofibrils of the muscle fiber fracture and the fragments retract and form a helical sheath around the central myofibrils of the same muscle fiber.