High-resolution DNA content analysis of microbiopsy samples in oral lichen planus.

OBJECTIVES: DNA aneuploidy has been reported to be a predictor of poor prognosis in both premalignant and malignant lesions. In oral lichen planus (OLP), this hypothesis remains to be proved. This study aimed to determine the rate of occurrence of DNA aneuploidy in patients with OLP by high-resolution DNA flow cytometry. METHODS: Patients with OLP were consecutively enrolled. Tissue samples were subdivided for formalin fixation and routine histological assessment and for immediate storage at -20°C for later DNA ploidy analysis, which was performed by DAPI staining of the extracted nuclei and excitation with a UV lamp. The DNA aneuploid sublines were characterized by the DNA Index. RESULTS: A DNA aneuploid status was observed in two of 77 patients with OLP (2.6%). When considering the clinical aspect of the OLP lesions, both DNA aneuploid cases had a reticular clinical aspect. CONCLUSIONS: DNA aneuploidy is an uncommon event in OLP and less frequent compared to other non-dysplastic and non-OLP oral potentially malignant disorders. The extremely low rate of DNA aneuploidy could represent an occasional finding or reflect the low rate of malignant transformation observed in patients with OLP even if the real prognostic value of DNA ploidy analysis in patients with OLP remains to be confirmed.

Congenital muscular dystrophies with defective glycosylation of dystroglycan: a population study.

BACKGROUND: Congenital muscular dystrophies (CMD) with reduced glycosylation of alpha-dystroglycan (alpha-DG) are a heterogeneous group of conditions associated with mutations in six genes encoding proven or putative glycosyltransferases. OBJECTIVES: The aim of the study was to establish the prevalence of mutations in the six genes in the Italian population and the spectrum of clinical and brain MRI findings. METHODS: As part of a multicentric study involving all the tertiary neuromuscular centers in Italy, FKRP, POMT1, POMT2, POMGnT1, fukutin, and LARGE were screened in 81 patients with CMD and alpha-DG reduction on muscle biopsy (n = 76) or with a phenotype suggestive of alpha-dystroglycanopathy but in whom a muscle biopsy was not available for alpha-DG immunostaining (n = 5). RESULTS: Homozygous and compound heterozygous mutations were detected in a total of 43/81 patients (53%), and included seven novel variants. Mutations in POMT1 were the most prevalent in our cohort (21%), followed by POMT2 (11%), POMGnT1 (10%), and FKRP (9%). One patient carried two heterozygous mutations in fukutin and one case harbored a new homozygous variant in LARGE. No clear-cut genotype-phenotype correlation could be observed with each gene, resulting in a wide spectrum of clinical phenotypes. The more severe phenotypes, however, appeared to be consistently associated with mutations predicted to result in a severe disruption of the respective genes. CONCLUSIONS: Our data broaden the clinical spectrum associated with mutations in glycosyltransferases and provide data on their prevalence in the Italian population.

POMT1 and POMT2 mutations in CMD patients: a multicentric Italian study.

Mutations in POMT1 and POMT2 genes were originally identified in Walker-Warburg syndrome (WWS) and subsequently reported in patients with milder phenotypes characterised by mental retardation with or without brain abnormalities and without ocular malformations. As part of a multicentric Italian study we screened the POMT1 and POMT2 genes in 61 congenital muscular dystrophy (CMD) patients with alpha-dystroglycan reduction on muscle biopsy and/or clinical and radiological findings suggestive of the known forms of CMD with alpha-dystroglycan deficiency. The aim of the study was to establish how frequently mutations in POMT1 and POMT2 occur in CMD patients in the Italian population and to evaluate the spectrum of associated phenotypes. Thirteen patients showed mutations in POMT1 and five harboured mutations in POMT2, accounting for a total of 20 different mutations, eight of which were novel (two in POMT1 and six in POMT2). Normal brain MRI associated with mental retardation and microcephaly was the most frequent finding in patients with mutations in POMT1 (six out of 13), but was also found in a patient with POMT2 mutations. Predominant cerebellar hypoplasia was also frequent both in patients with POMT1 (three out of 13) and POMT2 (three out of 5) mutations. A MEB phenotype with frontal cortical dysplasia and pons abnormalities was found in two patients with POMT1 and in one with POMT2 mutations, while a WWS phenotype was only found in a case with mutations in POMT1. Mutations causing frameshifts and stop codons were responsible for the more severe phenotypes. Our results provide further evidence that, as previously reported for FKRP, the array of mutations in POMT1 and POMT2 is ample and the spectrum of associated phenotypes is wider than initially thought.

A histochemical approach to the evaluation of the in vivo cytotoxicity of the nitrobutadienes (1E,3E)-1,4-bis(1-naphthyl)-2,3-dinitro-1,3-butadiene and methyl (2Z,4E)-2-methylsulfanyl-5-(1-naphthyl)-4-nitro-2,4-pentadienoate in mice liver and kidney.

Two new molecules (1E,3E)-1,4-bis(1-naphthyl)-2,3-dinitro-1,3-butadiene (1-Naph-DNB) and (2Z,4E)-2-methylsulfanyl-5-(1-naphthyl)-4-nitro-2,4-pentadienoate (1-Naph-NMCB) in previous studies showed interesting antiproliferative activity in vitro. Furthermore, toxicological tests and histological analysis provided promising results, in particular for 1-Naph-NMCB that displayed lower toxic activity both in terms of lethal effect and tissue damage of the main organs. Finally, studies of the antitumour activity in vivo confirmed the efficacy of both molecules, though with some differences in tumour selectivity and levels of activity. In this investigation the activities of some specific enzymes, acid phosphatase (AcPase), alkaline phosphatase (AlkPase), catalase (Cat), succinic dehydrogenase (SDH), glucose-6-phosphatase (G6Pase) and K+ p-nitrophenyl phosphatase (K+ pNPPase) were studied in the liver and kidney as histopathological biomarkers, to assess the effects of the two compounds in organs generally involved in the metabolism and excretion of different drugs. As oxidative stress may also develop as a consequence of the toxic effect of chemicals, reactive oxygen species (ROS) production was evaluated by a histochemical method. The results indicated that some enzyme activities and ROS expression changed in a dose-related manner. Nevertheless, neither in the liver nor in the kidney were dramatic toxic effects evident. By contrast, the variations of some enzyme activities (AlkPase, AcPase, Cat, K+ pNPPase) were interpreted as possible defensive mechanisms for tolerating high dosage of the compounds.

Preparation and evaluation of nanoparticles made of chitosan or N-trimethyl chitosan and a cisplatin-alginate complex.

In this work, nanoparticles with a negative or positive surface charge were prepared through electrostatic interaction of an anionic cisplatin-alginate complex with a cationic polyelectrolyte, namely chitosan or N-trimethyl chitosan (substitution degree of 85%). Statistical experimental design allowed the study of the influence of component amounts on the characteristics of nanoparticles. Mean particle diameter ranged from 180 nm to 350 nm. After 24 h, while the cisplatin-alginate complex released almost all the drug in saline-buffered solution at pH 7.4, approximately 40% w/w of total cisplatin was released from negative nanoparticles and roughly 50% w/w from positive ones. The same cumulative amounts of released drug were found after 48 h, with a progressive reduction to lower values up to 6 days. Drug loading of nanoparticles with a positive zeta potential (43 mV-60 mV) ranged from 13% w/w to 21% w/w and particle yield, referred to total polymers, was about 15% w/w (50% w/w if referred to cisplatin-alginate complex). Nanoparticles with a negative zeta potential (-34 mV) were obtained with a yield of 40% w/w and a drug loading of 18% w/w. These nanoparticles were the least active on all cell lines tested, while the cytotoxic activity of the positive nanoparticles was similar to or lower than that of cisplatin, probably depending on the combination of sizes and zeta potential values, on P388 murine and A2780 human cells. On A549 human cells, the nanoparticles with the smallest size and the lowest positive zeta potential were more active than cisplatin and showed a similar capability in inducing apoptosis in A2780 human cells. These results indicate that cisplatin complexes with polycarboxylate polymers can be transformed into cisplatin particulate carriers of high potential interest.

Somatostatin analogues, a series of tissue transglutaminase inducers, as a new tool for therapy of mesenchimal tumors of the gastrointestinal tract.

Imatinib, a tyrosine kinase inhibitor directed against the enzymatic domain of KIT protein, was found to produce dramatic clinical responses in metastatic gastrointestinal stromal tumors (GISTs). However, resistance usually develops thus determining treatment failure. The present study was performed to analyse the expression of somatostatin receptor (SSTR) subtypes, modulators of tissue transglutaminase, in a series of GISTs and leiomyosarcomas by immunohistochemistry to identify a new potential therapeutic target. Sixteen cases (8 males and 8 females, age range: 38-73; 11 GISTs, 4 leiomyosarcomas, 1 leiomyoma) were studied. Immunohistochemical detection of the relevant SSTRs was performed on paraffin-embedded tissue sections, stained with polyclonal antibodies directed against the five somatostatin receptor subtypes. We found 7 out of 16 (44%) tumors expressing all SSTRs and 14 out of 16 (87%) tumors positive for at least 3 subtypes. SSTR2A was the most represented subtype in the tumors studied, being expressed in approximately 70% of cases exhibiting an intense labeling in most of these cases. The significant expression of SSTRs shown in this series of GISTs and gastrointestinal leiomyosarcomas suggests a potential therapeutic target to be explored alone and/or in combination with other therapeutic agents in the setting of refractory GI stromal tumors.

Immunomodulatory properties of cefaclor: in vivo effect on cytokine release and lymphoproliferative response in rats.

The proper and coordinated response of the host immune system to bacterial infections is known to play a central role in the eradication of an infection. Therefore, the impact of antibiotics on both innate and acquired host immunity may be involved in the therapeutic outcome. The aim of this study was to evaluate the effects of the widely used cephalosporin cefaclor on some parameters of the immune system in ex vivo conditions. The results demonstrated that short-term (3 to 6 days) treatment with this antibiotic induced pleiotropic modification of rat spleen cells upon ex vivo stimulation with the polyclonal mitogen PHA, entailing increased lymphoproliferative responses, augmented IFN-gamma, IL-2 and IL-10 synthesis and decreased production of IL-4 and IL-6 in comparison to spleen cells from control rats. The mononuclear spleen cells of healthy rats released larger amounts of IFN-gamma and IL-2 in culture supernatants in response to polyclonal mitogenic stimulation with PHA compared to the spleens of the control rats receiving vehicle only. Simultaneously, the treatment with cefaclor augmented PHA-induced lymphoproliferative responses and reduced the synthesis of IL-4 and IL-6. These data depict a type 1 cytokine inducing and immunostimulatory pharmacological profile that, by activating the innate and acquired immune system, would be synergistic with cefaclor antibacterial activity.

Massive lower gastrointestinal hemorrhage in patients with portal hypertensive enteropathy: a report of two cases.

Mucosal and vascular changes in the lower gastrointestinal tract occur commonly in patients with portal hypertension. Portal enteropathy, however, is usually asymptomatic, though occasionally clinically significant for chronic gastrointestinal bleeding. Massive hemorrhage has only rarely been described and its management is controversial. Even though more effective non-operative treatments are now available, an emergency porta-systemic shunt procedure remains an important option for selected patients. We report on two cases of massive lower gastrointestinal bleeding from portal hypertensive enteropathy secondary to post-viral cirrhosis.

[Treatment of recto-urethral fistulas in Crohn's disease]. / Trattamento delle fistole retto-uretrali nel morbo di Crohn.

We report the thirteenth case of a rectourethral fistula in Crohn's disease. The patient, a 37 year-old-white male, had a 20 year history of intestinal Crohn's disease and had undergone numerous bowel resections. His symptoms were fecaluria, urorrhea and passing of urine from an orifice just outside the base of the scrotum. He had urinary infection and severe ileocolitis. He underwent a diagnostic evaluation that revealed a fistula comprising the membranous urethra, the rectum, the perineum and the scrotum. We performed medical therapy with metronidazole (20 mg/kg/day/12 months). We present in this article a review of the literature on the management of rectourethral fistulas in Crohn's disease. Surgeons have successfully used several approaches in the repair of this disorder, but no single procedure had proved to be best or even universally applicable. We emphasize, as the literature suggests, that management must be individualized. Medical therapy with metronidazole has an important role in a patient with rectourethral fistula and concomitant proctitis, ileocolitis, urinary sepsis and multiple previous surgeries.

Fibroblasts regulate the migration of MCF7 mammary carcinoma cells in hydrated collagen gel.

We have defined a tissue culture method suitable to study cell-cell interactions in an environmental set close to in vivo conditions. It consists of heterotypic cell populations mixed together inside a collagen gel in a chamber slide for a period of up to 14 days. When the three-dimensional system is saturated, cells will start to move on the plastic surface as monolayers surrounding the gel, with a characteristic speed depending on cell type. Usually fibroblasts move fast, while epithelial cells demonstrate a much lower pace of migration. At any given time gel contraction can be measured, and thus the rate of cell expansion, by knowing the distance from the edge of the gel to the leading edge of cell migration. By using this approach it was found that MCF7 mammary carcinoma cells display a great variety of morphologies following their mixture with different fibroblastic cell lines. In particular, when MCF7 cells were mixed with fibroblasts from human fetus, dog thymus and rat kidney, they migrated up to the leading edge of the fibroblastic front as isolated single cells or as cellular aggregates, many of which became necrotic in time, or took on an elongated morphology. Selective necrosis of MCF7 cells was also induced with serum concentration of 15% and 20% FCS, but only when they were mixed with fibroblasts. No necrosis was induced in MCF7 cells cultured alone. From these observations it is suggested that necrosis may sometimes favor the detachment and infiltration of resistant epithelial tumor cells by increasing their autonomous behaviour. Fibroblasts seem to be instrumental in regulating this process.

[Chronic idiopathic intestinal pseudo-obstruction: a clinical case and review of the literature]. / Pseudostruzione intestinale cronica idiopatica: caso clinico e revisione della letteratura.

The authors report a case characterized by impairment of the intestinal function clinically mimicking a mechanical obstruction in the absence of any occluding lesion of the gut lumen. Different pathological conditions, different clinical syndromes, instrumental approach for the diagnosis as well as medical and surgical treatment of pseudo-obstruction are discussed. Surgery has not been proven to be effective in the clinical management of CIIP for the irreversibility of the myopathic or neuropathic lesions. The most frequent causes of death are unnecessary surgery, ab ingestis pneumonia, cardiac arrest and malnutrition.

DNA damage induced by 7,12-dimethylbenz[a]anthracene in the liver and the mammary gland of rats exposed to polycyclic aromatic hydrocarbon enzyme inducers during perinatal life.

The long-lasting modulating effect induced by the prenatal or neonatal exposure to phenobarbital (PB) and aroclor on the genotoxic activity of 7,12-dimethylbenz[a]anthracene (DMBA) in female Sprague-Dawley rats was studied. The effect was measured as DNA damage evaluated in the liver and in the mammary gland of 55-day-old animals, 4 and 24 h after an i.g. injection of 80 mg/kg of DMBA. PB was given per os, i.g. or in drinking water to pregnant females and by i.g. only to neonates or in adult progeny. Aroclor was injected i.g. in prenatal and in neonatal life, and a second dose was given in adult life. Under these experimental conditions it was shown that DNA damage kinetics caused by DMBA are modulated by exposure to PB and, to a minor extent, by aroclor. The amount and persistence of DNA damage were highest when PB was administered to neonates. An average 2-fold increase in the elution constants (K) of DNA in the liver and the mammary gland was observed 4 h after DMBA treatment, as compared to uninduced animals. Repeated enzyme induction by PB seems to reduce DMBA genotoxicity, as shown by a decrease in DNA damage and persistence in the liver and mammary gland. The inducibility of the monooxygenase enzyme system in perinatal life favouring metabolic activation or inactivation of polycyclic aromatic hydrocarbons might be critical in determining individual susceptibility of adult progeny to chemical carcinogenesis by DMBA.

Modifying factors in prenatal carcinogenesis (review).

A description of the experimental approaches devised to control the growth of tumors induced by transplacental exposure to carcinogens is given. Due to the massive cell proliferation and differentiation taking place during embryogenesis, fetal tissues are believed to be privileged targets of neoplastic changes. As a consequence, trace amounts of environmental carcinogens capable of accumulating into the conceptuses may determine the appearance of tumors in the offspring, a possibility documented in several animal species including humans. Endogenous and exogenous factors counteracting this process have potential application as regulators of developmental carcinogenesis. Their identification is regarded as a means to chemoprevent pediatric tumors and can be instrumental in the analysis of the aetiopathogenesis of neoplastic phenotypes.

[The diagnosis of the cardiologic risk in patients destined for lung removal surgery]. / Diagnosi del rischio cardiologico in pazienti destinati a chirurgia di exeresi polmonare.

The expanding role of pulmonary surgery could be expected to lead to an increase in postoperative morbidity and mortality unless surgical risk factors are identified by means of a careful preoperative assessment. A careful and intelligent preoperative evaluation can reduce the morbidity and mortality after lung resection by ensuring the appropriate selection of patients and a rational choice of operative procedure. To evaluate the cardiologic risk in patients undergoing lung resection, 18 of them underwent a diagnostic protocol based on non invasive analysis including ECG, stress test with basal and exercise blood gas analysis, M- and B-mode and Doppler echocardiography. This protocol was able to identify possible cardiologic risk in the perioperative period without applying invasive techniques and with a good cost-benefits ratio.

Skin papillomas and other neoplasms induced by murine sarcoma viruses in mid-gestation-infected mice.

During extensive investigations on the effects of oncogenic retroviruses in developing rodents, the ability of MSV to mount a neoplastic response in CD-I Swiss mouse embryos was determined. By infecting the animals directly in utero at selected stages of post-implantation development, we detected a peculiar reaction of the embryonal tissues to certain MSVs: when mice were exposed to KiMSV at mid-gestation, the newborn developed characteristic tumors, in addition to mesenchymal cell sarcomas, not induced in fetuses and neonates. These included pulmonary alveologenic tumors and skin papillomas and were seen in mice infected on days 8 and 10 of pregnancy, roughly corresponding to 15 and 35 somites, respectively. To determine the specificity of these events, other 8- and 10-day-old embryos were infected with retroviruses of the same or different families. HaMSV and MoMSV also induced mesenchymomas and a low incidence of skin papillomas (10% and 15% compared to 40% in the KiMSV group) but not pulmonary tumors. In contrast, FBRMSV was inactive in this respect and only osteogenic sarcomas were detected in the offspring. Infecting the embryos on day 7 of pregnancy produced no tumors. Later infections (in 15-day-old fetuses and neonates) mainly induced mesenchymal sarcomas. No congenital malformations were detected in the embryos exposed to MSV during organogenesis, although some abortions and resorptions were seen.

Cystic angiomatosis (hamartous haemolymphagiomatosis) of bone. A clinicopathological study of three cases.

The clinical and radiological features in three cases of cystic angiomatosis of bone are reported. Although these features are generally diagnostic except from histiocytosis X, the definitive diagnosis must be established by a pathological study, preferably of a segment of an involved rib or fibula. The prognosis varies according to the type of clinical presentation-in particular upon whether the lesions are solely skeletal or whether there is extraskeletal visceral involvement. Whereas these last cases may often prove fatal, those with only skeletal involvement have a favourable prognosis: indeed, the cystic bone lesions may regress without any treatment, as occurred in some cases reported in the literature and in two of our three cases.