Hepatic expression of miR-122, miR-126, miR-136 and miR-181a and their correlation to histopathological and clinical characteristics of patients with hepatitis C.

It has been shown for hepatitis C virus (HCV) infection that host miRNAs contribute to the replication of the viral RNA genome. However, the clinical impact of these and many other cellular miRNAs on HCV in humans is still largely unclear. We therefore analysed the expression of miR-122, miR-126, miR-181a and miR-136 in HCV-infected patients. The study included liver biopsies of 65 patients infected with HCV of different genotypes (gt 1, gt 1a, gt 1b, gt 3 and gt 4) and nine noninfected individuals. Expression analysis of miRNAs was performed by qPCR, and they were analysed for differences between patient gender and age, genotypes, stage of fibrosis, grade of inflammation, serum level of liver enzymes, serum viral load, the presence of steatosis and mode of transmission. Different target prediction algorithms were used to search for targets of analyzed miRNAs. Statistical analysis revealed significant up-regulation of miR-136 and down-regulation of miR-126 and miR-181a in patients infected with HCV of different genotypes compared with noninfected individuals. The same expression pattern was observed in different stages and grades of liver disease. miR-122 was up-regulated in women relative to men and associated to portal inflammation, miR-122 and miR-126 correlated with serum HCV load and miR-136 and miR-122 correlated with the presence of steatosis. miR-126 and miR-136 were differentially expressed between different modes of HCV transmission. There were approximately 2000 different targets predicted for all four miRNAs and each of the analyzed miRNAs could be involved in more than a 100 different biochemical pathways. miR-122, miR-126, miR-136 and miR-181a have been shown to be involved in HCV infection with different genotypes. Their expression has been associated with the gender, stage and grade of liver disease, mode of transmission, serum HCV load and the presence of steatosis. Numerous target genes and biochemical pathways are predicted for each of the analyzed miRNAs. All these results suggest their role in HCV-infected liver disease.

Widespread vesiculobullous eruption in a 16-year-old male.

Bullous systemic lupus erythematosus (BSLE) is a rare but distinct disease, characterized by vesiculobullous skin eruptions and systemic lupus erythematosus (SLE). It can arise either before or after a diagnosis of SLE has been established. BSLE is characterized by a dermatitis herpetiformis-like histology and an autoimmunity to type VII collagen. It must be differentiated from other autoimmune vesiculobullous diseases such as epidermolysis bullosa acquisita, dermatitis herpetiformis, linear IgA disease, and bullous pemphigoid. A combination of clinical, histological, and immunofluorescence findings are necessary to establish a diagnosis of BSLE. We present a case of BSLE to illustrate and emphasize the need for an integrative diagnostic approach.

An uncommon case of chronic leg ulcers in an 80-year-old woman.

Ulcers of the lower extremities, particularly in individuals older than 65, are a common cause for visits to the dermatologist, primary-care physician, or vascular surgeon. There are many different causes of leg ulcers, among which vascular causes are the most frequent. Less commonly, other pathological processes result in leg ulcers. Unfortunately some of them are malignant. Large B-cell lymphoma, leg type, is a malignant lymphoma of intermediate behavior, occurring mostly on the legs in elderly patients. Usually it presents as erythematous or bluish-red nodules or tumors, but ulcerations are not uncommon. When faced with unusual and non-healing ulcers, the physician should also have in mind rarer but more lethal causes.

Expression of human telomerase catalytic protein in gallbladder carcinogenesis.

BACKGROUND: Telomerase catalytic subunit (hTERT) gene re-expression is a rate limiting step for the activity of telomerase, a key enzyme implicated in cellular immortalisation and transformation. AIMS: To determine the potential role of hTERT protein in gallbladder carcinogenesis. MATERIAL/METHODS: hTERT protein was analysed by means of immunohistochemistry in 89 gallbladder tissue samples: 16 normal epithelia, 14 reactive hyperplasias, 15 low grade dysplasias, 16 high grade dysplasias, and 28 adenocarcinomas. At least 200 nuclei were assessed for each slide and the mean number of positive signals for each nucleus was expressed as the hTERT index. RESULTS: The mean hTERT index increased progressively with the degree of gallbladder epithelial abnormalities: from 0.03 in normal epithelia, 0.04 in hyperplastic epithelia, 0.25 in low grade dysplasia, 0.82 in high grade dysplasia, to 0.93 in adenocarcinoma. Statistical analysis revealed that three different groups of gallbladder epithelial changes can be distinguished according to the number of hTERT signals for each nucleus: (1) normal and regenerative gallbladder epithelium, (2) low grade dysplasia, and (3) high grade dysplasia and adenocarcinoma (p < 0.001). CONCLUSIONS: The occasional presence of hTERT protein in normal and regenerative gallbladder mucosa reflects their regenerative capacity. Nevertheless, significantly higher hTERT indices in low and high grade dysplastic epithelia and in gallbladder adenocarcinomas are probably a consequence of hTERT re-expression--an early event in the multistep process of gallbladder carcinogenesis.

Acute fatty liver of pregnancy -- an underlying condition for herpes simplex type 2 fulminant hepatitis necessitating liver transplantation.

The infrequent occurrence of herpes simplex virus (HSV) hepatitis in healthy women in comparison with the high prevalence of HSV infections suggests that, in addition to deranged immunity, an underlying condition in the liver might be necessary to develop HSV hepatitis. We report the case of a 28-year-old pregnant woman in the 28 (th) week of gestation. Following HSV type 2 infection of the uterine cervix, acute liver failure developed, necessitating urgent liver transplantation. In addition to fulminant HSV type 2 hepatitis, the explanted liver also showed the histological features of acute fatty liver of pregnancy. The presented case suggests a possible pathogenetic role of acute fatty liver of pregnancy in the development of fulminant HSV hepatitis following recurrent infection with HSV in healthy pregnant women. We believe that early histopathological diagnosis, followed by specific antiviral treatment and liver transplantation in selected patients may improve the clinical outcome of otherwise almost uniformly fatal HSV hepatitis.

Human telomerase catalytic subunit gene re-expression is an early event in oral carcinogenesis.

AIMS: Detection of telomerase catalytic subunit (hTERT) mRNA has been used as a surrogate marker for estimation of telomerase activity. The exact role and timing of telomerase re-activation, a key enzyme implicated in cellular immortalization and transformation, in the multistep process of oral carcinogenesis is still unknown. The aim was to test the hypothesis that (i) quantitative rather than qualitative differences exist in the level of hTERT mRNA expression between normal oral mucosa, different grades of oral epithelial abnormalities and squamous cell carcinomas of the oral cavity, and that (ii) hTERT gene re-expression is an important, probably early event in oral carcinogenesis. METHODS AND RESULTS: The relative quantity of hTERT mRNA was analysed in 45 frozen oral epithelia representing different morphological stages of oral carcinogenesis classified according to the Ljubljana classification and in 37 oral squamous cell carcinomas, using a commercially available LightCycler Telo TAGGG hTERT Quantification kit. hTERT mRNA was not detected in normal or reactive hyperplastic oral epithelia, but was present in 43% of atypical hyperplasias (premalignant lesions), 60% of intraepithelial carcinomas and 68% of oral squamous cell carcinomas. Statistical analysis revealed two groups of oral epithelial changes, with significant differences in the levels of hTERT mRNA expression: 1, normal and reactive hyperplastic oral epithelium, and 2, atypical hyperplasia, intraepithelial carcinomas and squamous cell carcinomas. CONCLUSION: These data suggest that hTERT gene re-expression represents an early event in the multistep process of oral carcinogenesis, already detectable at the stage of precancerous oral epithelial changes. Nevertheless, other genetic aberrations appear to be necessary for progression of oral epithelial abnormalities towards invasive squamous cell carcinoma.

Treatment of chronic hepatitis C: our experience.

BACKGROUND/AIMS: While an optimal treatment of chronic hepatitis C has not yet been established, it has been demonstrated that the interferon alpha/ribavirin combination is more effective than interferon alpha monotherapy. METHODOLOGY: One hundred and forty-three patients with chronic hepatitis C received the following treatment: eighty patients an 18-month monotherapy (3-month follow-up) and sixty-three patients a 12-month combined therapy (6-month follow-up). Therapeutic efficacy and adverse effects were compared. RESULTS: In 80 patients in the monotherapy group, complete response was achieved in 49.2%. This was reduced to 27.5% three months after therapy. Significant differences were observed in HCV 3 genotype where complete response was achieved in 12 out of 14 patients (p=0.01). With the combined therapy administered to 63 patients, complete response was achieved in 54.5%. This was reduced to 43.2% after 6 months of follow-up. Among the responders or partial responders, significant differences were observed with regard to age (p=0.0047) and subtype 1b (p=0.012). Comparing the groups of naive patients and relapsers, a statistically significant difference (p=0.027) was found in therapeutic efficacy. CONCLUSIONS: In the treatment of chronic hepatitis C, combined therapy proved more effective than monotherapy. This is, however, not yet a satisfactory solution.

Inflammatory myofibroblastic tumour of paranasal sinuses with fatal outcome: reactive lesion or tumour?

Inflammatory myofibroblastic tumours (IMTs) are clinicopathologically distinctive but biologically controversial entities, which have been described in the lungs, abdomen, retroperitoneum, and extremities, but rarely affect the head and neck region. IMT usually follows a benign clinical course after radical excision, but invasive, locally recurrent, and metastatic forms of abdominal and mediastinal IMT have also been described. This report describes a case of IMT of the paranasal sinuses with a fatal outcome. A 22 year old woman was admitted to hospital as a result of epistaxis. Computed tomography scan and magnetic resonance imaging showed an expansive process in the paranasal sinuses, extending into the nasal cavity, orbita, and endocranium. The tumour progressed despite several surgical procedures. Radiotherapy, corticosteroids, and chemotherapy were unsuccessful, and the patient died four years after diagnosis, as a result of extensive intracranial spread of the tumour. This is the first known case of an IMT of the head and neck region with a fatal outcome. It shows that the aggressive behaviour of IMTs is not limited to abdominal and mediastinal locations, and supports recent observations that at least a subset of IMTs represents true neoplasia rather than reactive myofibroblastic proliferation.

Human papillomavirus DNA in oral squamous cell carcinomas and normal oral mucosa.

To elucidate the putative etiologic role of human papillomaviruses (HPV) in oral carcinogenesis, a comparative study was carried out on 62 tissue specimens of oral squamous cell carcinoma (OSCC) and on 62 specimens of histologically normal oral mucosa obtained from the individuals who matched the subjects with OSCC in age, gender, localization of obtained tissue specimens, drinking and smoking habits. Internal control amplification showed that amplifiable DNA was recovered from 59/62 and 61/62 tissue samples of OSCC and normal oral mucosa, respectively. The amplification with two different HPV L1 and one HPV E6 consensus primer sets showed the presence of the HPV DNA genotypes 16, 33, 58 in 5/59 (8.4%) OSCC specimens and HPV genotypes 11, 16, 31, 68 in 4/61 (6.6%) tissue samples of normal oral mucosa tested. In the study in which a comparative examination of the presence of HPV DNA was for the first time performed on the tissue samples of the patients with OSCC and the age- and gender-matched control subjects there was no significant difference in the prevalence of HPV DNA among both study groups. Our results suggest that occasional findings of HPV DNA in OSCC tissue specimens may be the result of an incidental HPV colonization of oral mucosa, rather than of viral infection, and that HPVs play a limited role in the etiopathogenesis of the majority of OSCC.

Proliferative and apoptotic activity in hepatocellular carcinoma and surrounding non-neoplastic liver tissue.

UNLABELLED: The aim of the study was to determine the proliferative and apoptotic activity of neoplastic and non-neoplastic hepatocytes, to ascertain whether there was a correlation between the histopathological characteristics of hepatocellular carcinoma (HCC) and its proliferative or apoptotic activity. METHODS: One tumour sample and one sample of non-neoplastic liver from 16 patients with HCC were analysed. The proliferative activity was established by immunohistochemical staining against PCNA (proliferating cell nuclear antigen) and Ki-67. Apoptotic activity was determined by morphological and TUNEL methods. Bcl-2 immunoreactivity was analysed. RESULTS: A positive correlation between PCNA and Ki-67 proliferative indexes was found in HCC (p < 0.01). The PCNA index was 0.21% +/- 0.80% (Mean +/- SD) in non-neoplastic liver and 7.41% +/- 8.22% in HCC, while the Ki-67 index was 0.19% +/- 0.26% in non-neoplastic liver and 9.67% +/- 7.70% in HCC. The differences between HCC and non-neoplastic liver were significant (p < 0.05). The PCNA index differed significantly between different HCC grades (p < 0.05). In HCC and non-neoplastic liver samples, apoptotic indexes (AI) assessed morphologically were higher than Al determined by the TUNEL method. Differences in Al (irrespective of the method used) between different HCC grades were not significant. Bcl-2 staining was positive in one non-neoplastic liver sample (6.3%) and in 4 HCC samples (25%). CONCLUSIONS: PCNA and Ki-67 were useful for proliferative activity assessment of hepatocytes. There were no differences in apoptotic activity between HCC and non-neoplastic tissue, so it seems that uncontrolled tumour cell division plays an important role in HCC growth. In the regulation of the apoptotic process in HCC, Bcl-2 could be important.

Quantitative measurement of telomerase catalytic subunit (hTERT) mRNA in laryngeal squamous cell carcinomas.

We tested 30 laryngeal squamous cell carcinomas (LSCCs) and 30 matched control laryngeal samples from the same patients for the presence of human telomerase catalytic subunit (hTERT) mRNA by using the Roche LightCycler Telo TAGGG hTERT Quantification kit. The hTERT index was calculated to express the relative quantity levels of hTERT mRNA. hTERT mRNA was detectable in 10 out of 30 (33%) laryngeal tissues covered by normal and/or reactively hyperplastic laryngeal epithelium and 23 out of 30 LSCCs (77%). The mean hTERT indices were 0.15 for control non-cancerous laryngeal samples, 0.57 for grade I, 2.35 for grade II and 3.72 for grade III LSCCs. LSCCs without detectable hTERT mRNA (23%) tended to have lower grades of disease. No correlation was found between the levels of hTERT mRNA and tumour size or locoregional lymph node status. We believe that hTERT mRNA in normal and/or reactively hyperplastic laryngeal epithelium originates from the stem cells and corresponds to the self-renewal capacity of the squamous epithelium. However, the greater quantity of h TERT mRNA in LSCCs is the result of telomerase reactivation in the process of laryngeal carcinogenesis.

Role of lipids in the progression of renal disease in systemic lupus erythematosus patients.

Systemic lupus erythematosus (SLE) is an autoimmune connective tissue disease marked by immune-complex mediated lesions in small blood vessels of various organs, especially the kidneys, although other factors may also be implicated in the pathogenesis of the disease. This article focuses on the role of lipids in the progression of glomerular, vascular and tubulo-interstitial lesions in two patients with lupus nephritis associated with pronounced hyper- and dyslipidemia. The pathogenesis of progressive glomerulosclerosis in both patients appears to be multifactorial. In addition to immune complex mediated lupus glomerulonephritis, progressively active in the first patient, severe nephrotic-range persistent proteinuria, arterial hypertension associated with hyperfiltration and hyperperfusion injuries and, to a minor extent, hyper- and dyslipidemia were observed. Immunological and non-immunological factors were shown to contribute to the development of tubulo-interstitial lesions. In both patients, in addition to local immune deposits, prominent tubulo-interstitial lipid deposits were probably causally related to both hyperlipidemia and the increased permeability of the glomerular filtration barrier. Tubular lesions were highlighted by intracytoplasmic lipid droplets as well as small cleft-like spaces found to be impacted in the tubular lumina. They were seen to penetrate tubular epithelial cells and eventually lodge in the interstitium, surrounded by mononuclear cell infiltrates and foam cells. In both patients, hypertensive angiopathy and extraglomerular vascular immune deposits were demonstrated. In addition, in the second patient, arteriolar and small arterial hyaline was found at the age of 28 years to be full of lipids and calcium precipitates, suggesting a peripheral atherosclerosis-like process which never occurs as a natural age-related condition. In conclusion, all parts of the nephron may be involved in the pathogenetic process causally related or influenced by hyper- or dyslipidemia. Associated either with endothelial cell injury and consequent insudation of lipids in the vascular walls, glomerular filtration barrier injury with hyperfiltration, or tubulo-interstitial lipid deposition, the mechanism of tissue damage by lipids in all parts of the nephron shares similarities with the pathogenesis of systemic atherosclerosis.

Laryngeal granuloma: characteristics of the covering epithelium.

The purpose of the present study was to evaluate the biological behaviour of the marginal epithelium, that proliferates and eventually covers laryngeal granulomas, and to reveal the applicability of the recently re-introduced Ljubljana classification when reporting reactive epithelial hyperplastic lesions. A retrospective clinical and histomorphological analysis was performed on 149 laryngeal granuloma biopsies. Epithelial changes were classified according to the Ljubljana classification into normal epithelium; simple, abnormal, or atypical hyperplasia; and carcinoma in situ. Atrophic epithelium, not evaluated separately in the Ljubljana classification, was additionally assessed. Simple hyperplasia was found in 98 cases (65.8 per cent), abnormal hyperplasia in seven (4.7 per cent), atrophic epithelium in 24 (16.1 per cent), and normal squamous epithelium in 20 (13.4 per cent). Atypical hyperplasia and carcinoma in situ were not observed. The results of our study clearly showed that the proliferation of the covering epithelium mostly in the form of simple hyperplasia, is entirely reactive and therefore reversible. No epithelial hyperplastic lesions were found that were previously described to be associated with an increased risk of malignant alteration, namely atypical hyperplasia and carcinoma in situ. However, since an initial growth of an invasive malignant neoplasm might macroscopically imitate the appearance of laryngeal granuloma, a histological examination in all aetiological forms of laryngeal granulomas is required. By clearly discerning the benign nature of epithelial changes in laryngeal granulomas, the recently re-evaluated and further formulated Ljubljana classification may also influence the clinical handling of patients.

Infantile HIV encephalopathy associated with cerebral and cerebellar telangiectases.

We describe a paediatric case of HIV encephalopathy associated with cerebral and cerebellar telangiectases. Although immunohistochemistry failed to show HIV in the walls of dilated blood vessels, or in their vicinity, brain capillary telangiectases might be an additional complication indirectly related to paediatric HIV infection.

p53 protein overexpression in laryngeal squamous cell papillomas.

We retrospectively investigated p53 protein immunoreactivity in 103 laryngeal squamous cell papillomas (LP) previously revealed to be human papillomavirus type 6 or 11 positive by in situ hybridization and/or the polymerase chain reaction. 21 LP failed to show any detectable level of p53 protein reactivity. In 45 cases only occasional strongly positive cells were observed in almost the whole thickness of the epithelium. In 26 LP, p53 protein immunoreactivity was found to be almost exclusively restricted to the basal epithelial cells. Finally, in 11 cases, basal cell layer immunoreactivity was accompanied by aggregates of p53-positive cells in the lower two thirds of the epithelium. This staining pattern was found predominantly in LP with atypical hyperplasia. We think that the observed patterns of p53 immunoreactivity in a majority of cases are a result of immunohistochemical detection of the stabilized wild-type p53 protein rather than the mutated p53 protein.

Langerhans and other immunocompetent cells in vocal cord epithelial hyperplastic lesions of patients with chronic laryngitis.

The aim of the study was to evaluate the intraepithelial and stromal density of Langerhans cells and lymphoid infiltrate in different stages of carcinogenesis in vocal cord biopsies of 24 randomly selected patients with chronic laryngitis. The Langerhans and lymphoid cells were counted using immunolabelling with antibodies against CD1a, S100, CD3, CD20, and CD68 on paraffin-embedded sections of 24 archival laryngeal vocal cord mucosa biopsy specimens, 6 classified as simple, 7 as abnormal, and 11 as atypical epithelial hyperplasia. Results were statistically evaluated using the Kruskal-Wallis and Wilcoxon sign rank tests. The mean number of Langerhans cells and T lymphocytes per mm2 of cross-sectioned epithelium was found to increase from simple to atypical hyperplasia. There were statistically significant differences in Langerhans cell density between atypical hyperplasia and each of the other 2 grades, simple and abnormal hyperplasia, with p < 0.05. Our study suggests the involvement of immune mechanisms, particularly cell mediated, during laryngeal carcinogenesis and the possibility that the assessment of Langerhans cell density might be of prognostic significance.

Langerhans cells in human papillomaviruses types 6/11 associated laryngeal papillomas.

Some studies have shown a reduced density of Langerhans cells (LCs) within epithelium infected by human papillomaviruses (HPV) types 16/18. However, data on a correlation between HPV types 6/11 infection and LCs have been missing. To solve this problem, we analysed 24 biopsy specimens of laryngeal papillomas, selected randomly, 20 multiple and 4 solitary. The presence of HPV 6 and 11 was proven by polymerase chain reaction (PCR) using 2 different sets of primers in 23 biopsy specimens. Abnormalities of the covering stratified squamous epithelium were graded according to the Kambic-Gale classification. LCs were immunohistochemically labelled with 2 different antibodies, CD1a and S100. Quantitative analysis was performed to determine the density of LC per mm2 in different grades of epithelial abnormalities covering laryngeal papillomas. Although no statistically significant differences in the mean number of LCs per mm2 of the cross-sectioned epithelium covering laryngeal papillomas were observed comparing simple, abnormal and atypical hyperplasia groups, the mean number of LCs per mm2 in laryngeal papillomas associated with HPV types 6/11 infection substantially exceeded that of the vocal cord surface epithelium in patients with chronic laryngitis.

Correlation of histomorphological criteria used in different classifications of epithelial hyperplastic lesions of the larynx.

Different classifications of epithelial hyperplastic lesions of the larynx were proposed, but none of them has been generally accepted. The basic distinction among these gradings is evaluation of carcinoma in situ as a precancerosis or a distinct and separate entity. In the present study, atypical hyperplasia and carcinoma in situ are evaluated according to the proposed histomorphological criteria of the Kambic-Lenart classification. In an attempt to separate more objectively the histomorphological differences between these 2 lesions, in addition to traditional light microscopical examination, we also performed semiquantitative analysis with statistical evaluation using the Wilcoxon signed rank test. These results revealed a significant morphological and statistical difference comparing atypical hyperplasia and carcinoma in situ on the basis of the following criteria: abnormal mitotic figures (p = 0.005), mitotic activity (p = 0.014), nuclear pleomorphism (p = 0.006), cellular atypia (p = 0.005), dyskeratosis (p = 0.008), and stromal infiltration (p = 0.015). These results confirm our view that atypical hyperplasia and carcinoma in situ are 2 consecutive but different entities in the process of carcinonogenesis.