Quantitation of cytokine gene expression by real time PCR in bovine milk and colostrum cells from cows immunized with a bovine rotavirus VP6 experimental vaccine.

In a previous work, VP6 recombinant protein was produced using baculovirus system and it was evaluated in a colostrum-deprived calf model. This vaccine was able to protect calves against viral challenge without inducing neutralizing antibodies (NAb), suggesting that another immunological effectors were involved in the protection observed. In this work, groups of cows (n=4) were immunized in the last third of gestation with a bovine rotavirus (BRV) experimental vaccine and with a VP6 subunit vaccine. At birth, colostrums from vaccinated and non-vaccinated cows were processed and viable colostral mononuclear cells were obtained. With the purpose of determining the cytokine patterns generated by cells from immune secretions (colostrums and milk), a relative quantification by real time PCR was standardized. Quantitative real time PCR (qPCR) was used to determine transcript levels of IL-4, IL-6, IL-10, IL-12, IFN-γ and IFN-α from these cells. Colostral and milk mononuclear cells expressed a different cytokine transcript expression pattern regarding the vaccine used. These results demonstrated that the colostral cellular population was active and could exert its action influencing the final immune response.

Molecular characterization of chicken infectious anemia virus circulating in Argentina during 2007.

Chicken infectious anemia virus (CAV) is a worldwide-distributed infectious agent that affects commercial poultry. Although this agent was first detected in Argentina in 1994, no further studies on CAV in this country were reported after that. The recent increased occurrence of clinical cases of immunosuppression that could be caused by CAV has prompted this study. Our results confirmed that CAV is still circulating in commercial flocks in Argentina. Phylogenetic analysis focusing on the VP1 nucleotide sequence showed that all Argentinean isolates grouped together in a cluster, sharing a high similarity (> 97%) with genotype B reference strains. However, Argentinean isolates were distantly related to other strains commonly used for vaccination in this country, such as Del-Ros and Cux-1. Sequence analysis of predicted VP1 peptides showed that most of the Argentinean isolates have a glutamine residue at positions 139 and 144, suggesting that these isolates might have a reduced spread in cell culture compared with Cux-1. In addition, a particular amino acid substitution at position 290 is present in all studied Argentinean isolates, as well as in several VP1 sequences from Malaysia, Australia, and Japan isolates. Our results indicate that it is possible to typify CAV strains by comparison of VPI nucleotide sequences alone because the same tree topology was obtained when using the whole genome sequence. The molecular analysis of native strains sheds light into the epidemiology of CAV in Argentinean flocks. In addition, this analysis could be considered in future control strategies focused not only on breeders but on broilers and layer flocks.

Relationships between blood pressure, anthropometric characteristics and blood lipids in high- and low-altitude populations from Central Asia.

We studied the relationships between blood pressure, anthropometric characteristics and blood lipids in 72 low altitude (LA) Uighurs (600 m), 91 LA-Kirghizs (900 m), 117 medium altitude (MA) Kazakhs (2100 m) and 94 high altitude (HA) Kirghizs (3200 m). All subjects were male and had a similar age (p = ns, ANOVA; range for all 374 subjects: 18-66 yr). Body weight (Wt), body mass index (BM1) and the sum of four skinfolds (4SF) were significantly lower in HA-Kirghizs than the remaining groups (p < 0.0005, p < 0.0005 and p < 0.05 respectively, ANOVA). However, no difference was found in body fat distribution as detected by waist:hip circumference (WHR) and triceps:subscapular skinfold ratios (TSR; p = ns, ANOVA). Stage 1 hypertension was detected in 18% of LA-Uighurs, 2% of LA-Kirghizs, 4% of MA-Kazakhs and 1% of HA-Kirghizs; stage 2 hypertension was detected in 2% of LA-Uighurs and none of the remaining groups; no subject had stage 3 hypertension (The Joint National Committee on Prevention. Detection, Evaluation and Treatment of High Blood Pressure 1997). Blood cholesterol (CH) and triglycerides (TG) did not differ between groups (p = ns, ANOVA). The relationships between systolic (SBP) or diastolic (DBP) blood pressure and age, Wt, BMI, 4SF, WHR, TSR, CH and TG were independent from altitude (p = ns, ANCOVA). In the pooled sample (n = 374), age explained 1 and 3% of SBP (p < 0.05) and DBP (p < 0.005) variance respectively, Wt was the best predictor of SBP and DBP explaining 11 and 10% of their variance respectively (p < 0.0001) and CH explained 5% of DBP variance (p < 0.0001). In conclusion, hypertension is more frequent in LA- than MA- and HA-subjects from Central Asia. However, anthropometric characteristics and blood lipids do similarly contribute to explain blood pressure in these subjects.

Taste sensitivity to PTC and thyroid function (FT4 and TSH) in high- and low-altitude Kirghiz populations in the Pamir.

PTC taste sensitivity distribution, determined using the Harris and Kalmus method, and analysis of thyroid activity using FT4 and TSH hormone assays were studied in a sample of 108 high-altitude subjects (3200 m) and in 90 lowlanders (900 m) from two different regions of Kirghizstan (Central Asia). All subjects were healthy Kirghiz males. In agreement with other available data on Kirghiz populations, a higher nontaster frequency was found in the high-altitude subjects. Furthermore, the results of our study indicate no association between thyroid function, PTC sensitivity, and age in both samples. No difference in thyroid function indicators was noted between highlanders and lowlanders.

Sclerosing hemangioma of the lung: a case report.

A well circumscribed nodular mass discovered on routine chest ray examination, was excised from the left lung interlobar fissure of a 67 years old woman. Histologically the lesion corresponded to a typical "so called sclerosing hemangioma" according to Liebow and Hubbell's originally description. Because of the uncertain histogenesis we utilized histochemistry and immunohistochemistry to support the hypothesis of an epithelial origin.

Ascites apheresis, concentration and reinfusion for the treatment of massive or refractory ascites in cirrhosis.

A new method for ascites recirculation, consisting of a cellulose diacetate filter to remove substances with molecular weight > or = 300,000, cell debris and bacteria, followed by the concentration of ascitic fluid prior to i.v. infusion, was used 24 times in 19 patients with cirrhosis and massive or refractory ascites. The amount of ascites removed was 7.67 +/- 0.49 l, which was reduced to 407 +/- 37 ml. The procedure took 367 +/- 22 min to complete. No statistically significant changes in liver function tests, coagulative parameters, platelet count or natremia were found. The activity of coagulation and fibrinolytic systems was further assessed in six patients. No changes suggesting an activation of intravascular coagulation and/or primary fibrinolysis were disclosed. An asymptomatic fall in mean arterial pressure (from 88.6 +/- 2.6 to 80.3 +/- 3.0 mmHg; p = 0.02) occurred after paracentesis and was still present 48 h after ascites reinfusion. Plasma renin activity significantly decreased at the end of the procedure, but was not associated with a proportional reduction of plasma aldosterone concentrations. Both variables returned to baseline values 48 h later. A significant increase in the glomerular filtration rate occurred just after the end of the procedure (from 50.4 +/- 9.1 to 73.1 +/- 23.5 ml/min; p < 0.05) and subsided 48 h later. In contrast, no significant changes in diuresis and renal sodium excretion were found. Complications due to volume overload and sepsis did not occur; in one case, spontaneous bacterial peritonitis developed 3 days after the procedure.(ABSTRACT TRUNCATED AT 250 WORDS)

Factors that affect keratotomy depth.

The authors investigated nine factors which can affect the depth of incisions performed during refractive keratotomy: (1) vertical vs oblique-cutting edge of the knife blade, (2) direction of cutting, (3) cutting velocity, (4) American vs Russian technique, (5) intraocular pressure (IOP), (6) initial vs final incisions, (7) sharpness of knife blade, (8) single vs double footplate, and (9) square vs double-edged blade. These variables were examined independently, performing at least 40 incisions for each experimental parameter studied. The depth of the resulting incisions was measured histologically using the micrometer eyepiece. The average and the standard deviation were calculated. The paired Student's t-test was used to establish significant differences between the two conditions investigated for each parameter. Factors that were demonstrated to increase significantly the depth of the incisions included: the vertical-cutting edge, the triple-edged diamond knife, the sharpness of the knife, and the single foot knife. High velocity in performing the incisions and, to a lesser extent, low IOP were the main factors that induced irregularity in depth.

Basal cell carcinoma arising in pemphigus vulgaris of the lower eyelid.

A 59-year-old woman on continuous treatment with corticosteroids and immunosuppressive drugs for pemphigus vulgaris of 14 year's duration presented with an enlarging nodular lesion on the right lower eyelid. The neoplasm, diagnosed as basal cell carcinoma, was excised conserving the lid margin, and eyelid reconstruction was done with a rotated cheek flap. Wound healing proceeded without complications in spite of immunosuppressive treatment, and the final cosmetic and functional result was satisfactory.

[Effect of CDP-choline on senile mental deterioration. Multicenter experience on 237 cases]. / Effetto della CDP-colina sul decadimento mentale senile. Esperienza policentrica su 237 casi.

The efficacy of CDP-choline (1000 mg/die) administered for two 21-day treatment cycles, with a one-week wash-out period between them, was evaluated in out and in-patients suffering from mild to moderate brain aging. The study was performed on 237 fully evaluable patients with the use of the reduced geriatric scale of Plutchik and al., for clinical evaluation of the symptomatology. The clinical data obtained demonstrate that treatment with CDP-choline is able to determine an improvement of symptomatology since the 1st cycle of therapy (p less than 0.001), and a further improvement in the 2nd cycle (p less than 0.001). Particularly, the therapeutic effect of the 1st cycle is persistent in the intermediate wash-out period (suspension of treatment) with a further decrease, of symptomatology regarding some items of Plutchik's scale (p less than 0.01). Finally, treatment with CDP-choline 1000 mg/die for two 21-day cycles in 237 patients suffering from brain aging determined a statistically significant improvement of the cognitive and behavioural parameters taken into consideration: independence/autonomous life; human relations/social life; interest and attentive capacity; individual behaviour. Therefore citicoline is confirmed as a valid therapeutic remedy for the clinical, functional and social recovery of these patients.

Different methylation of oestrogen receptor DNA in human breast carcinomas with and without oestrogen receptor.

The methylation of the human oestrogen receptor (ER) gene was analysed by restriction enzymes in normal and neoplastic human breast tissues and cell lines. CCGG sequences in regions inside the gene, which are methylated both in normal breast and in tissues that are not the target of the oestrogen, are hypomethylated in 30% of tumours, both ER+ and ER- carcinomas. Moreover, 5' sequences of the gene, which are hypomethylated in normal breast and not in tissues not the target of oestrogen, are methylated to a lower degree in ER+ carcinomas, whereas they are methylated to a greater degree in ER- carcinomas. However, the same region is equally hypomethylated in both ER+ and ER- cancer cell lines. Our results indicate that in breast carcinomas ER DNA methylation is deranged, and in cancer cell lines is different from that observed in primary tumours. Furthermore, the abnormal methylation in the 5' end seems to be related to abnormal expression, namely diffuse hypomethylation in carcinomas with high ER content and hypermethylation in carcinomas without ER. These findings support our previous hypothesis that DNA methylation could be involved in the control of ER gene expression and demonstrate that abnormal ER gene methylation is a typical feature of breast cancers.

The methylation pattern in the 5' end of the human estrogen receptor gene is tissue specific and related to the degree of gene expression.

Restriction enzyme mapping has been used to study the relationship between methylation at the 5' end of the human estrogen receptor gene (ER) and its various levels of expression in estrogen target tissues, like endometrium, placenta and breast. The methylation patterns were compared with that found in white blood cells. A cluster of CpG sites, including the transcription start site, was undermethylated in all examined tissues. In front and downstream of this area, a 1 Kb region was undermethylated in endometrium where the ER gene is highly expressed, methylated to some extent in placenta and breast, which express the gene to a low degree, and greatly methylated in white blood cells which are thought not to express this gene. These data provide evidence that in the 5' end of the human ER DNA, CpG methylation is tissue-specific, and related to the level of the ER gene expression.

Hypomethylation of the human HLA-DR alpha gene in breast carcinomas and autologous metastases.

The methylation pattern of the human HLA-DR alpha gene was analyzed in normal breast tissues, breast primary tumors and lymphonodal metastases isolated from patients carrying breast carcinomas. In breast adenomas and also in normal tissues (including breast, muscle, brain, sperm and T- and B-lymphocytes), the HLA-DR alpha gene is hypermethylated at the CCGG and GCGC sites. In all tissues studied, the only constantly unmethylated region is located in the 5' portion of the gene, near the promoter sequence. Further, the results indicate that the HLA-DR alpha gene is hypomethylated in carcinomas and in the relative metastatic lymph nodes. It is suggested that hypomethylation of the human HLA-DR alpha gene could be proposed as a molecular marker of malignant breast tumors.

Abnormal methylation of estrogen receptor gene and reduced estrogen receptor RNA levels in human endometrial carcinomas.

Demethylation of specific sites or restricted genomic regions has been reported to correlate with gene activation and also with carcinogenesis. As abnormal expression of Estrogen Receptor (ER) could be involved in the genesis or progression of tumors in estrogen target tissues, the methylation of ER gene has been compared in 8 endometrial carcinomas and 29 normal endometria. In order to look for a correlation between methylation and expression, levels of ER RNA were also measured. While the 5' region of ER gene was found to be demethylated in both normal and carcinomatous tissues, there was demethylation of some specific sites in the internal part of the gene only in the carcinomas examined. In addition, in the carcinomatous tissues the levels of ER RNA were low, indicating that an increase of ER gene hypomethylation does not raise, and even may reduce, the ER expression in endometrium. The abnormal undermethylation observed in ER gene appears to be unrelated to general DNA hypomethylation which is frequently present in neoplastic tissues; nor has it been found in ER DNA isolated from breast carcinomas. These data strongly support the hypothesis that such a methylation is specifically related to endometrial transformation and therefore it can be considered an additional marker of this disease.

Effect of intravenous and intraperivenous injections of sclerosants (sodium tetradecyl sulfate and hydroxy polyethoxy dodecan) on the rat femoral vein.

The sclerosant effect of injected tetradecyl sulfate of sodium (STS) and hydroxy polyethoxy dodecan (HPD) was studied in the rat femoral vein. Intravenous (i.v.) and intravenous plus perivenous (i.v. + p.v.) injections of both sclerosants and physiologic saline were compared as to vein lumen occlusion, fibrosis, phlogosis, and damage to the artery and surrounding nervous and muscular tissues. The study was carried out in 30 rats treated by STS, in 30 treated by HPD, and 15 animals were injected with saline. The neurovascular bundle and adjacent muscle were removed at 48 h, 7 and 30 days and examined histologically. I.v. injections of STS produced a solid occlusion of the vein in a significant number of cases, after 30 days (P less than 0.01). A statistically significant number of solid occlusions of the femoral vein resulted after i.v. + p.v. injection of STS and HPD, at 48 h, 7 and 30 days (P less than 0.05; P less than 0.01). There was no significant difference between STS and HPD after i.v. + p.v. injection. After i.v. + p.v. we recorded a marked inflammation of muscle with signs of focal necrosis, at 48 h and 7 days. Our study indicated that i.v. + p.v. injection of STS and HPD provided a high degree of efficacy as regards vein occlusion. On the other hand, i.v. + p.v. injection induced a severe inflammation and necrosis of the tissues surrounding the sclerosed vein. Extrapolating our results to the endoscopic sclerotherapy for esophageal variceal bleeding, we conclude that paravariceal injection of sclerosants is a dangerous procedure, even though efficacious to reduce variceal hemorrhage, owing to the high risk of iatrogenic ulcers and esophageal perforation caused by muscular and mucosal necrosis.