Human skeletal muscle-derived stem/progenitor cells modified with connexin-43 prevent arrhythmia in rat post-infarction hearts and influence gene expression in the myocardium.

Stem cell therapy in combination with genetic modification (e.g., transfection with the coding sequence for the connexion 43 gene, GJA1) may solve the problems associated with the occurrence of additional (secondary) stimulation in the post-infarcted heart (arrhythmia). Human skeletal muscle-derived stem/progenitor cells (SkMDS/PCs) were transfected with the pCiNeo-GJA1 plasmid at an efficiency of approximately 96%. Gene overexpression was assessed using qPCR, and subsequent analysis revealed that GJA1 expression increased more than 40-fold in SkMDS/PCs transfected with the appropriate coding sequence (SkMDS/PCsCX43) compared to that of the 'native' SkMDS/PCs control (SkMDS/PCsWT). Enhanced (4-fold) protein expression of connexin-43 was also confirmed by Western immunoblotting. Furthermore, using the arrhythmic score, we demonstrated the positive effects of SkMDS/PCsCX43 cell intervention in reducing additional secondary stimulations in rat post-infarcted hearts compared with that of wild-type cell delivery. Selected gene responses (Kcnq1, Cacna1c, Ncx1, Serca2a, and Tgfb1) showed significantly altered expression profiles in the rat myocardium upon intervention with SkMDS/PCsCX43. The genetic modification of human skeletal muscle-derived stem/progenitor cells with connexin-43 prevented the pro-arrhythmic effects of myogenic implanted stem cells on the host myocardium and positively influenced myocardial gene expression profiles in respect to myocardium conductivity.

Acroangiodermatitis in a leg amputee using a suction-socket prosthesis: clinical, histological as well as HHV-8 and CD34 immunohistochemical study.

Acroangiodermatitis (AAD) is a rare, vascular phenomenon of unclear pathogenesis. Itchy, lichenoid, purple/violaceous/yellowish/brownish papules/nodules, plaques/patches mainly on lower limbs occasionally evolve into verrucous lesions and recurrent painful ulcerations. Elevated vein and capillary pressure due to the sub-atmospheric suspension system seems to be the triggering factor for angioproliferation in the amputation stump. A middle-aged male amputee, a suction-socket prosthesis user, showing combined clinical, histological and immunohistochemical (HHV-8 negative; CD34 and CD31 expressed in endothelial, but not perivascular, cells) features of AAD is presented. Dermatologists, orthopedic surgeons, pathomorphologists, but also prosthesis makers and amputees themselves, should be aware of AAD as suction-socket prostheses become increasingly popular.

Heligmosmoides polygyrus fourth stages induce protection against DSS-induced colitis and change opioid expression in the intestine.

Primary exposure of mice to the nematode Heligmosomoides polygyrus infection reduces inflammation in an experimental model of colitis. The aim of the present investigation was to evaluate whether the reduced inflammation provoked by H. polygyrus L4 larvae in BALB/c mice treated with dextran sulphate sodium is associated with changed expression of opioids in the small intestine and colon. Colitis was induced by 5% Dextran sulphate sodium (DSS) oral administration for 3 days before oral infection with 200 infective larvae (L3) H. polygyrus until the end of the experiment, 6 days post-infection. Clinical disease symptoms were monitored daily. The expressions of proopiomelanocortin POMC1, MOR1 (Oprm1) - opioid receptor and ß-endorphin were determined by RT-PCR, Western blot and immunoassay, respectively, in the colon and small intestine of mice. RT-PCR analysis of colon tissues showed up-regulation of the expression of POMC and MOR1 opioid-dependent genes in mice with DSS-induced colitis. H. polygyrus L4 larvae inhibited DSS-induced colitis symptoms that were correlated with increased IL-1ß, TNF-α, IL-6, myeloperoxidase (MPO) concentration, macrophages infiltration and MOR1, POMC and ß-endorphin increased expression in the small intestine and inhibition of those in the colon.

Pouchitis may increase the risk of dysplasia after restorative proctocolectomy in patients with ulcerative colitis.

AIM: Dysplasia of the pouch mucosa after restorative proctocolectomy is rare. The aim of this study was to establish whether there is a correlation between pouchitis and dysplasia. METHOD: A group of 276 patients treated for ulcerative colitis by restorative proctocolectomy between 1984 and 2009 was analysed. The presence or absence of pouchitis and dysplasia within the pouch was evaluated. RESULTS: Inflammation was diagnosed in 66 (23.9%) patients, low-grade dysplasia in five (1.8%), high-grade dysplasia in three (1.1%), and cancer in one patient (0.4%). The prevalence of low-grade dysplasia was significantly higher in patients with inflammation than in those without (P < 0.04). High-grade dysplasia was significantly more frequent in pouchitis than in non-inflamed pouches (P < 0.01). Logistic regression analysis suggested that the occurrence of mucosal inflammation increased the risk of low grade dysplasia. CONCLUSION: Patients with chronic pouchitis are at risk of dysplasia and require surveillance of the pouch.

IL-6, TNF-alpha, IL-1-beta arterial wall expression is independent of serum concentration in patients sustaining primary or secondary open vascular reconstructions.

AIM: The aim of this study was to assess morphological and immunohistochemical differences in arterial wall inflammation during primary and secondary vascular reconstructions. METHODS: Forty patients with lower extremities ischemia underwent vascular reconstruction: Group I included patients undergoing primary vascular reconstruction with 25 minor arterial reconstructive procedures; Group II included patients undergoing secondary reconstruction due to limb ischemia after at least 12 months since first operation (15). Immunohistochemical expression of IL-1-BETA,IL-6,and TNF-alpha in arterial wall was correlated with serum concentration. RESULTS: No differences were observed in IL-1-beta, IL-6, TNF-alpha expression among groups (P=0.06, P=0.13, P=0.9). Cytokines expression was not accompanied by the growth of its serum concentrations (P=0.8, P=0.9). IL-6 serum concentration depended on the degree of limb ischemia and was very high in case of critical limb ischemia (P=0.006). Arterial morphology was comparable among the groups. CONCLUSION: IL-6, IL-1-BETA, TNF-alpha expression in the arterial wall is comparable in primary and secondary lesions. Serum concentrations of IL-6, IL-1-BETA, TNF-alpha are independent of its arterial wall expression.

Campylobacter protein oxidation influences epithelial cell invasion or intracellular survival as well as intestinal tract colonization in chickens.

The Dsb family of redox proteins catalyzes disulfide bond formation and isomerization. Since mutations in dsb genes change the conformation and stability of many extracytoplasmic proteins, and since many virulence factors of pathogenic bacteria are extracytoplasmic, inactivation of dsb genes often results in pathogen attenuation. This study investigated the role of 2 membrane-bound oxidoreductases, DsbB and DsbI, in the Campylobacter jejuni oxidative Dsb pathway. Campylobacter mutants, lacking DsbB or DsbI or both, were constructed by allelic replacement and used in the human intestinal epithelial T84 cell line for the gentamicin protection assay (invasion assay) and chicken colonization experiments. In C. coli strain 23/1, the inactivation of the dsbB or dsbI gene separately did not significantly affect the colonization process. However, simultaneous disruption of both membrane-bound oxidoreductase genes significantly decreased the strain's ability to colonize chicken intestines. Moreover, C. jejuni strain 81-176 with mutated dsbB or dsbI genes showed reduced invasion/intracellular survival abilities. No cells of the double mutants (dsbB⁻ dsbI⁻) of C. jejuni 81-176 were recovered from human cells after 3 h of invasion.

The influence of elastic components of the venous wall on the biomechanical properties of different veins used for arterial reconstruction.

OBJECTIVE: To evaluate the relationship between the biomechanical properties and the structure of elastic components in different veins used for vascular reconstruction. DESIGN: In vitro experimental study. MATERIAL AND METHODS: Groups of 30 samples of incompetent saphenous veins (rSV), competent saphenous veins (cSV) and femoral veins (FVs) were compared following immunohistochemical staining for the presence of collagen types I, III and IV and elastin. The percentage area of transverse section of veins occupied by each type of collagen and elastin was measured using a computer-image-analysis system connected to a microscope. For all three groups of veins, the storage modulus, E', and the loss modulus, E'', were measured with a mechanical analyser, DMA-242, and changes in the function of temperature and frequency, and duration of exposure to the applied force were determined. RESULTS: The rSV showed the highest percentage share of collagen I and the lowest percentage share of collagen IV. These samples also showed the greatest expression of elastin and the highest elastin to collagen ratio. The rSV were also found to have the highest E' and E'', and during the long-term exposure achieved maximum stiffness in the least time as compared to cSV and FV. CONCLUSION: The histological structure directly influences the biomechanical properties of venous wall with rSV showing least compliance and cSV the greatest compliance.

Single brain metastasis from a minute, well differentiated, but invading beyond the tunica muscularis mucosa rectal carcinoid.

We report the case of a 48-year-old woman with small-sized (< 10 mm), highly differentiated, aggressive rectal carcinoid, who developed a solitary distant metastasis to the brain. The primary lesion, initially removed by conventional polypectomy, invaded the mucosa/tunica muscularis mucosa and had positive resection margins. Afterwards, an assessment of 5-hydroxyindoloacetic acid (5-HIAA) 24 h urine excretion revealed a significantly increased level. Thus, a partial rectal resection was performed. Because of constantly elevated carcinoid markers: serum chromogranin A (CGA) and 5-HIAA, a somatostatin receptor scintigraphy was performed, which disclosed a focus of pathological marker accumulation in the left frontal area. The pathological finding after neurosurgical excision was meningioma. An unexpected normalization of the biochemical markers prompted us to verify this diagnosis. The final histopathological report was a well-differentiated neuroendocrine brain metastasis. Our case shows that in well differentiated, of diameter < 10 mm rectal carcinoids, an invasion even beyond the mucosa/tunica muscularis mucosa seems to be an independent factor predicting a malignant metastatic potential of these tumors. Hence, in such cases, behind the endoscopic submucosal resection with ligation device a more radical surgery should be considered. Additionally, a systematic CGA and 5-HIAA follow-up assessment and whole body somatostatin receptor scintigraphy, if necessary, are required.

Synthesis, surface modifications, and size-sorting of mixed nickel-zinc ferrite colloidal magnetic nanoparticles.

We report on the spontaneous covalent growth of monomolecular adlayers on mixed nickel-zinc nanoferrite colloidal suspensions (ferrofluids). Synthesized nanoparticles were subjected to surface modification by means of acid chloride chemistry, leading to the formation of covalent bonds between the hydroxy groups at the nanoparticle surface and the acid chloride molecules. This procedure can be easily tailored to allow for the formation of adlayers containing both hydrophobic and hydrophilic regions stacked at predetermined distances from the magnetic core, and also providing the nanoferrites with functional carboxy groups capable of further modifications with, for example, drug molecules. Here, fluorophore aminopyrene molecules were bound to such modified nanoferrites through amide bonds. We also used the same chemistry to modify the surface with covalently bound long-chain palmitoyl moieties, and for comparison we also modified the nanoferrite surface by simple adsorption of oleic acid. Both procedures made the surface highly hydrophobic. These hydrophobic colloids were subsequently spread on an aqueous surface to form Langmuir monolayers with different characteristics. Moreover, since uniformity of size is crucial in a number of applications, we propose an efficient way of sorting the magnetic nanoparticles by size in their colloidal suspension. The suspension is centrifuged at increasing rotational speed and the fractions are collected after each run. The mean size of nanoferrite in each fraction was measured by the powder X-ray diffraction (PXRD) technique.

Ghrelin ovarian cell expression in patients with polycystic ovary syndrome: an immunohistochemical evaluation.

INTRODUCTION: The influence of ghrelin on different organs has been studied recently, e.g. in the regulation of pituitary hormone release, regulation of energy homeostasis, glucose metabolism and insulin secretion, cell proliferation, and reproductive function. However, the etiology of polycystic ovary syndrome has not been fully explained. The aim of our study was to estimate the presence of ghrelin in polycystic ovaries cells and evaluation of the relationship between ghrelin occurrence and cells proliferation. METHODS: In the present work we have compared ten polycystic ovaries with ovaries without pathology as the control group. We used immunohistochemical method to detect ghrelin. The cells proliferation was evaluated by Ki 67 proliferation index. RESULTS: Ghrelin immunostaining was demonstrated in cytoplasm of ovarian secondary interstitial cells and in atretic corpus luteum. The cell nuclei were ghrelin positive in granulosa, theca layers of follicular cyst in both groups as well as in luteal cells of young corpus luteum in healthy ovaries. Ki 67 immunostaining was observed in granulosa and theca layers of follicular cyst in polycystic and healthy ovaries. CONCLUSIONS: It is possible that local ghrelin expression plays an important role in the direct control of ovarian development and function and ghrelin may participate in patomechanism of PCOS.

Quantitative evaluation of immuno- and histochemical reaction intensity by spatial visualization techniques.

The aim of this study was an application of spatial visualization techniques for quantitative measurements of immuno- and histochemical reactions. For a quantitative histochemical study, specimens, collected from patients with chronic gastritis, were stained with paS/AB, while for immunohistochemical evaluation, specimens were used, collected from patients with chronic parathyroiditis and were analyzed with Ki-67 proliferation marker and apoptosis bcl-2 protein. The new technique permitted to obtain quantitative objective results. Statistical cluster analysis of those results indicated small groups of cases for reevaluation and supported the final diagnosis.

[Accuracy of ultrasound-guided fine-needle aspiration biopsy in diagnosis of changes in thyroid nodular lesions]. / Skutecznosc celowanej biopsji aspiracyjnej cienkoiglowej w diagnostyce zmian guzowatych tarczycy.

The aim of this study was to compare the results of US-FNAB with definitive histological examination of thyroid nodular lesions. 590 patients who underwent surgery were reviewed (473 females, 117 males, ranging in age from 9 to 81 years, average 36 years). Histological evaluation of cytologically diagnosed benign nodules revealed nodular goiter in 407 cases (91.5%), Hashimoto's thyroiditis in 2 (0.4%), follicular adenoma in 31 (7%), papillary carcinoma in 2 (0.4%) and follicular carcinoma in 3 (0.7%). In the cytological group of follicular nodule (n = 71) histological diagnoses included: nodular goiter in 11 cases (15.5%), follicular adenoma in 36 (50.7%), papillary carcinoma in 2 (2.8%), follicular carcinoma in 20 (28.2%). The diagnosis of papillary carcinoma (n = 65) was confirmed histologically in 59 cases (90.8%), in the remaining 6 cases Hashimoto's thyroiditis and medullary carcinoma were diagnosed. In the cases diagnosed cytologically as medullary carcinoma (n = 8) histological diagnoses included: medullary carcinoma in 7 cases (87.5%). The cytological diagnosis of anaplastic carcinoma (n = 1) was confirmed histologically. These results support the value of US-FNAB in the diagnostics of thyroid neoplasms. US-FNAB performance was as follows: sensitivity 78%, specificity 97%, accuracy 92%, 2.3% of false positive and 6.1% of false negative results.

[Primary malignant melanoma of the larynx]. / Pierwotny czerniak zlosliwy krtani.

Primary malignant melanoma of the larynx is very rare. The case of larynx malignant melanoma in 71-years old men is presented. Hoarseness was the main complaint that encouraged the patient to visit a doctor. The patient was heavy smoker, he had been smoking 40 cigarettes for the last 50 years. He used to drink about 200 ml of pure alcohol per day. In the direct laryngoscopy gluish had cyanotic tumor, extending to the true vocal cord, ventricle, ventricle fold and subglottic space was seen. Enlarged cervical lymph nodes, laterally to the main vessels were found both in palpation and in US examination. Total laryngectomy and neck dissection was performed. The histological examination of surgical specimen revealed; Malignant melanoma. Metastases in the lymph nodes. Neoplastic infiltration of the nodal capsule. The patient was admitted to the complementary radiotherapy. At the moment the follow up in outpatient. The article contains the review of literature concerning the diagnosis, treatment and prognosis in cases of laryngeal malignant melanoma.

Endoscopical and histological aspects of inflammatory changes in J-pouch mucosa.

The aim of the study was endoscopic and histological evaluation of mucosal inflammatory changes in the ileal reservoir after restorative proctocolectomy with ileoanal anastomosis (IPAA) because of FAP (27 patients) and ulcerative colitis (34 patients). Endoscopic symptoms of pouchitis (edema, granularity, contact bleeding, loss of vascular pattern, mucus, erosions) were found in 13 patients (4 were operated on because of FAP, 9 because of UC). A statistically significant difference (p < 0.0001) was found between endoscopic inflammatory index in UC (4.0 +/- 0.5) and in FAP (1.00 +/- 0.00). The endoscopic changes were correlated with the Moskowitz histological classification, in 12 acute score and in 6 chronic score were above 4 points. In 18 patients histological diagnosis of pouchitis was made (5 in FAP, 13 in UC). Acute pouchitis occurred in 12 patients and chronic in 6. In one patient histological changes resembled Crohn's disease. Regular endoscopic control with histological examination and verification of inflammatory lesions in the ileal reservoir is necessary.

Identification and mutation analysis of DOC-1R, a DOC-1 growth suppressor-related gene.

The tumor suppressor gene MEN1 and several oncogenes including CCND1/cyclin D1/PRAD1 map to chromosome 11q13. However, molecular and cytogenetic analysis suggests the presence of a second tumor suppressor locus at this chromosome region. We have identified a novel gene from chromosome 11q13, which encodes a protein of 126 amino acids sharing an overall 57% identity with the p12(DOC-1) protein encoded by the DOC-1 gene, the human homolog of hamster putative tumor suppressor doc-1 (deleted in oral cancer-1). We therefore designated the novel gene as DOC-1R for DOC-1-related. The cytogenetic location was confirmed by chromosome fluorescent in situ hybridization. Northern blot analysis indicated that it was expressed in all the tissues examined. DOC-1R protein showed heterogeneous subcellular localization. RT-PCR-SSCP analysis failed to detect deleterious mutations of the DOC-1R transcript in four premalignant oral keratinocyte lines and 20 different cancer cell lines from tumor types which frequently harbor LOH at chromosome 11q13.

Mutational and expression analysis of the p73 gene in melanoma cell lines.

A novel p53-related gene, p73, was recently isolated and cytogenetically mapped to chromosome region 1p36. Functionally, p73 expression induces p21waf and suppresses tumor cell growth. We mapped p73 using radiation hybrids and localized the gene to an interval that putatively harbors a melanoma tumor suppressor locus. We then analyzed p73 transcripts from 24 melanoma cell lines using reverse transcription-PCR/single strand conformation polymorphism and identified nine polymorphic sequence changes (three novel and six previously published polymorphisms); furthermore, we found evidence of biallelic transcription in our cell lines. However, we did not detect any deleterious mutations. These data suggest that the p73 gene is unlikely to be essential in melanoma tumorigenesis.

[Retrospective analysis of selected tumor markers (p53, PCNA, Ki67; DNA ploidy) and ultrastructure in patients with larynx carcinomas]. / Retrospektive Analyse ausgewählter Tumormarker (p53, PCNA, Ki67; DNA-Ploidie) und der Ultrastruktur bei Kranken mit Larynxkarzinomen.

A comparison was made of the staining intensities of selected immunohistochemical proliferating antigens (p53, PCNA, Ki67), DNA flow-cytometry and ultrastructures of neoplastic cells from 120 cases of laryngeal cancers. Clinically very advanced tumors were in the majority (T3, 43%; T4, 18%). A 5-grade scale was adopted to evaluate the level of immunohistochemical staining of the carcinoma cell nuclei. Positive staining was obtained in 70% for p53, 57% for Ki67 and 80% for PCNA. Sixty-two percent of the cases were DNA-diploid and 38% DNA-aneuploid. The DNA-diploid carcinomas were accompanied by enlargement of the cell nuclei, preservation of wide margins of nuclear heterochromatin, enlargements of the nuclear areas and increases in the number of nuclei. In the aneuploid-polyploid cancers the nuclei had a substantial polymorphism, with large cleaved nuclei showing significant variations in size and having a nuclear envelope. A frequent finding was euchromatization of chromatin. Dense chromatin appeared in the form of small clumps spread over the whole area of these irregular nuclei. Enlargement and activation of nucleoli were found. There was a positive (chi-square) correlation between T- and N-stage and immunohistochemical staining. There was also a positive correlation in staining intensity between p53, Ki67 and PCNA markers and strong correlation between these markers for proliferative activity and the degree of aggressiveness of a tumor.

The level of MAP kinase activity in the stomach stump in rats after subtotal gastrectomy.

MAPK (Mitogen-Activated Protein Kinase) is one of the elements of kinase cascades (MAPK, MEK-MAP kinase, kinase, Raf-1, Ras) regulating cellular proliferation and differentiation processes. It seems that the changes in its number and activity may be the factor having influence on carcinogenesis. In some human carcinomas a significant increase of its activity is observed, in others a decrease of its activity is described. Our research aimed at the evaluation of the dynamics of precancerous and cancerous changes in the stomach stump in rats after the experimental, partial stomach resection. Apart from histological and ultrastructural examination we also determined the activity of the sub-unit p42 MAP kinase. The material comprised segments of gastric mucosa of the stomach stump of 15 rats after subtotal gastrectomy. Part of the rats after the procedure were administered carcinogen orally (MNNG). On the histological and ultrastructural examination we used routine methods, the activity of MAP kinase was determined by western-blotting method with the use of IgG against MAPK p42, Santa Cruz #154). In 8 examined rats we observed the increase of MAP kinase activity. We established probable correlation (without statistical analysis, regarding miserly material) between the increase of MAPK activity and histological and ultrastructural changes. Among three cases diagnosed as adenoma tubulare in two we observed the increase of MAPK activity. A clear increase of this kinase was also present in the stomach stump of a rat, which was diagnosed as adenocarcinoma. On the basis of our research carried so far we think that the increase of the MAPK activity may be one of the causes of the neoplasm development. It seems important to obtain the confirmation of our results and to establish a possible usefulness of MAPK activity determination as a prognostic indicator in case of the neoplasm of stomach stump.

[Sarcoma neurogenes of the larynx]. / Sarcoma neurogenes krtani.

Malignant nonepithelial neoplasms of the larynx are rare, and tumors of mesenchymal origin constitute 4-6% of all malignant neoplasms of the larynx. Sarcoma neurogenes vel malignant schwannoma make up 10% of all soft tissue sarcomas. In this paper a case of the extremely rare malignant schwannoma of the larynx is presented. In world literature we have met four case reports describing tumors of this histological morphology and location.

Ultrastructural changes in the small bowel epithelium after total gastrectomy.

The purpose of the study was to analyze ultrastructural changes in the small bowel mucosa in patients after total gastrectomy. We studied mucosal specimens obtained from 25 patients during control gastroscopy. The specimens were routinely processed for examination in transmission electron microscopy. Early after the operation (up to 6 months) we observed marked inflammatory reaction, disordered architecture of the small bowel mucosa epithelium, the presence of dysplasia-like changes and foci of dysplasia. Later on the structure of the mucosa returned to normality. Only a few dysplastic changes were seen. No relationship was found between altered epithelial structure and type of operation. In conclusion, the epithelium of the small bowel does not transform to a gastric type epithelium.