Study on prevalence and bacterial etiology of mastitis, and effects of subclinical mastitis and stage of lactation on SCC in dairy goats in Egypt.

In Egypt, inadequate information on prevalence and epidemiology of caprine mastitis is available. This study was designed to investigate prevalence and etiological agents of caprine mastitis and assess the efficacy of somatic cell count (SCC) as marker of subclinical mastitis (SCM) in dairy goats. This study was carried out on 249 randomly selected lactating goats in different lactation stages and examined clinically. Of these animals, 477 milk samples were aseptically collected and screened for bacterial carriage. SCC was assessed in 234 apparently normal milk samples, and SCC ≥ 106 cells/ml was indicator for SCM. Prevalence of clinical mastitis (CM) was 33.73% and 16.87% at animal and udder-half levels, respectively. SCM was 52.56% in the apparently healthy halves. Culture results proved single infection in 49.69% of samples, mixed infection in 23.9% of samples, and 26.41% of samples were negative. Coagulase negative staphylococci (CNS) were the most predominant bacteria (58.75%), then Staphylococcus aureus (S. aureus) (24.375%), and Streptococci (1.875%) were the least. No significant difference was recorded between mean of SCC in bacteriologically positive and negative samples, neither in those with SCC ≤ 106 nor with SCC ≥ 106 cells/ml both in middle and late lactation stages. Besides, the percentage of animals harboring SCC ≥ 106 cells/ml and negative for bacteriology in late lactation stage was 3 times (28.57%) more than in midlactation (9.3%). We can assume that SCC is not proper indicator for intra-mammary inflammation (IMI) in goats, and bacteriological examination remains more efficient, despites being time consuming and expensive.

Induction TPF followed by concomitant treatment versus concomitant treatment alone in locally advanced head and neck cancer. A phase II-III trial.

BACKGROUND: Platinum-based chemoradiation (CCRT) is the standard treatment for Locally Advanced Head and Neck Squamous-Cell Carcinoma (LAHNSCC). Cetuximab/RT (CET/RT) is an alternative treatment option to CCRT. The efficacy of induction chemotherapy (IC) followed by chemoradiation compared to chemoradiation alone has not been demonstrated in randomized clinical trials. The goals of this phase II-III trial were to assess: (i) the overall survival (OS) of IC versus no-induction (no-IC) and (ii) the Grade 3-4 in-field mucosal toxicity of CCRT versus CET/RT. The present paper focuses on the analysis of efficacy. MATERIALS AND METHODS: Patients with LAHNSCC were randomized to receive concomitant treatment alone [CCRT (Arm A1) or CET/RT (Arm A2)], or three cycles of induction docetaxel/cisplatin/5 fluorouracil (TPF) followed by CCRT (Arm B1) or followed by CET/RT (Arm B2). The superiority hypothesis of OS comparison of IC versus no-IC (Arms B1 + B2 versus A1 + A2) required 204 deaths to detect an absolute 3-year OS difference of 12% (HR 0.675, with 80% power at two-sided 5% significance level). RESULTS: 414 out of 421 patients were finally analyzed: 206 in the IC and 208 in the no-IC arm. Six patients were excluded because of major violation and one because of metastatic disease at diagnosis. With a median follow-up of 44.8 months, OS was significantly higher in the IC arm (HR 0.74; 95% CI 0.56-0.97; P = 0.031). Complete Responses (P = 0.0028), Progression Free Survival (P = 0.013) and the Loco-regional Control (P = 0.036) were also significantly higher in the IC arm. Compliance to concomitant treatments was not affected by induction TPF. CONCLUSIONS: IC followed by concomitant treatment improved the outcome of patients with LAHNSCC without compromising compliance to the concomitant treatments. The degree of the benefit of IC could be different according to the type of the subsequent concomitant strategy. CLINICAL TRIAL NUMBER: NCT01086826, www.clinicaltrials.gov.

Ambulatory blood pressure monitoring in the elderly: features and perspectives.

Aging leads to a multitude of changes in the cardiovascular system that include a rise in blood pressure. Age-related changes in blood pressure are mainly attributable to an increase in systolic blood pressure, generally associated with a slight decrease diastolic blood pressure. This leads to a widening in pulse pressure. Ambulatory blood pressure monitoring is a useful tool to understand these processes and to refine cardiovascular risk assessment. In the light of emerging data in this area, we reviewed the main features of ambulatory blood pressure in elderly and discussed the evidence showing that ambulatory blood pressure is superior to clinic blood pressure to reflect the true pattern of blood pressure over time. Furthermore, we discussed the role of weight control obtained by fitness programs to prevent an excessive rise in blood pressure with age. A thorough understanding of these concepts is of paramount importance and has therapeutic implications in the growing population of elderly subjects with increased blood pressure.

The treatment of the large periradicular endodontic injury.

AIM: Periradicular lesions of endodontic origin are characterized by polymicrobial infections, part of which appear to play a crucial role in the facultative anaerobic bacterical species. In literature there is a strong disagreement about the choice of treatment in large periradicular lesions of endodontic origin: some authors propose the orthograde root canal therapy, others surgical therapy with apicectomia, retrograde filling of the cavity and review instrument. The purpose of this study was to demonstrate the effectiveness of orthograde endodontic treatment in case of periapical lesions of endodontic origin of dimensions larger than 20 mm. METHODS: It was evaluated a sample of 60 cases, ages between 18 and 70 years, 32 men and 28 women. The cases have been treated by orthograde endodontic. Were included mono and pluriradicular teeth with periapical lesion of endodontic origin primary or secondary at endodontic incongruous treatment, with dimensions larger than 20 mm. The sample was divided into Group A: 19 cases in which was possible to complete the root canal therapy in the same event; Group B: 41 cases in which there was drainage. Dressing was applied with pure calcium hydroxide, which was renewed every 10 days for a maximum of 30, was eventually completed the endodontic therapy. RESULTS: Group A: 13 out of 19 cases showed healing at 5 years. Of the remaining 6, there were three failures, a crown-root fracture, missed two follow-up. At 10 years of the 13 successes, 2 cases showed relapse. Group B: 41 cases, later reduced to 30 we had 19 successes in 5 years. Of the remaining 11: 3 crown-root fractures, 2 missed the follow-up, 6 failures. At 10 years of the 19 successes, two were lost because of fracture, one for a relapse. Discussion. The results show the importance of drainage, which can affect the apical seal and therefore the success of endodontic therapy, but allows decompression of the periradicular lesion and symptoms regression. The use of calcium hydroxide in the intermediary dressings allows the neutralization of acidic compounds, alkaline phosphatase activation creating a significant development of the antibacterial action. Proper instrumentation and cleansing of root canals allows the reduction of over one thousand times the bacterial load. The coronal seal has, through the adhesive techniques of restorative materials, a crucial role in closing the doors of entry the bacterial contamination of treated root canals. CONCLUSION: The endodontic therapy by orthograde is considered primary therapeutic choice in case of large endodontic lesions, given the success at rate both 5 (Group A 68,41%, Group B 63,33%) and 10 years (Group A 57.88%, Group B 53.32%).

DNAJC17 is localized in nuclear speckles and interacts with splicing machinery components.

DNAJC17 is a heat shock protein (HSP40) family member, identified in mouse as susceptibility gene for congenital hypothyroidism. DNAJC17 knockout mouse embryos die prior to implantation. In humans, germline homozygous mutations in DNAJC17 have been found in syndromic retinal dystrophy patients, while heterozygous mutations represent candidate pathogenic events for myeloproliferative disorders. Despite widespread expression and involvement in human diseases, DNAJC17 function is still poorly understood. Herein, we have investigated its function through high-throughput transcriptomic and proteomic approaches. DNAJC17-depleted cells transcriptome highlighted genes involved in general functional categories, mainly related to gene expression. Conversely, DNAJC17 interactome can be classified in very specific functional networks, with the most enriched one including proteins involved in splicing. Furthermore, several splicing-related interactors, were independently validated by co-immunoprecipitation and in vivo co-localization. Accordingly, co-localization of DNAJC17 with SC35, a marker of nuclear speckles, further supported its interaction with spliceosomal components. Lastly, DNAJC17 up-regulation enhanced splicing efficiency of minigene reporter in live cells, while its knockdown induced perturbations of splicing efficiency at whole genome level, as demonstrated by specific analysis of RNAseq data. In conclusion, our study strongly suggests a role of DNAJC17 in splicing-related processes and provides support to its recognized essential function in early development.

Association of insulin receptor substrate 1 with simian virus 40 large T antigen.

Mouse embryo cells expressing a wild-type number of insulin-like growth factor I receptors (IGF-IR) (W cells) can be transformed either by simian virus 40 large T antigen (SV40 T) or by overexpressed insulin receptor substrate 1 (IRS-1), singly transfected. Neither SV40 T antigen nor IRS-1, individually, can transform mouse embryo cells with a targeted disruption of the IGF-IR genes (R- cells). However, cotransfection of SV40 T antigen and IRS-1 does transform R- cells. In this study, using different antibodies and different cell lines, we found that SV40 T antigen and IRS-1 are coprecipitated from cell lysates in a specific fashion, regardless of whether the lysates are immunoprecipitated with an antibody to SV40 T antigen or an antibody to IRS-1. The same antibody to SV40 T antigen, however, fails to coprecipitate another substrate of IGF-IR, the transforming protein Shc, and two other signal-transducing molecules, Grb2 and Sos. Finally, an SV40 T antigen lacking the amino-terminal 250 amino acids fails to coprecipitate IRS-1 and also fails to transform R- cells overexpressing mouse IRS-1. These experiments indicate that IRS-1 associates with SV40 T antigen and that this association plays a critical role in the combined ability of these proteins to transform R- cells. This finding is discussed in light of the crucial role of the IGF-IR in the establishment and maintenance of the transformed phenotype.

A functional insulin-like growth factor I receptor is required for the mitogenic and transforming activities of the epidermal growth factor receptor.

When wild-type mouse embryo cells are stably transfected with a plasmid constitutively overexpressing the epidermal growth factor (EGF) receptor (EGFR), the resulting cells can grow in serum-free medium supplemented solely with EGF. Supplementation with EGF also induces in these cells the transformed phenotype (growth in soft agar). However, when the same EGFR expression plasmid is introduced and overexpressed in cells derived from littermate embryos in which the insulin-like growth factor I (IGF-I) receptor genes have been disrupted by homologous recombination, the resulting cells are unable to grow or to be transformed by the addition of EGF. Reintroduction into these cells (null for the IGF-I receptor) of a wild-type (but not of a mutant) IGF-I receptor restores EGF-mediated growth and transformation. Our results indicate that at least in mouse embryo fibroblasts, the EGFR requires the presence of a functional IGF-I receptor for its mitogenic and transforming activities.

A soluble insulin-like growth factor I receptor that induces apoptosis of tumor cells in vivo and inhibits tumorigenesis.

By a frame-shift mutation, we have engineered a human IGF-I receptor (IGF-IR) cDNA that produces a receptor 486 amino acids long (plus the 30 amino acids of the signal peptide). This receptor, which we have designated as 486/STOP, is partially secreted into the medium of cells in culture and markedly inhibits the autophosphorylation of the endogenous IGF-IRs as well as the activation of the signaling pathway. The 486/STOP receptor acts as a strong dominant negative for several growth functions: (a) it inhibits the growth of cells in monolayers; (b) it inhibits the growth of transformed cells in soft agar; (c) it induces extensive apoptosis in vivo; and (d) it inhibits tumorigenesis in syngeneic rats. This is the first demonstration that a dominant negative of the IGF-IR can induce massive apoptosis of tumor cells in vivo.

Protective effect of the insulin-like growth factor I receptor on apoptosis induced by okadaic acid.

Okadaic acid (OKA), a potent inhibitor of serine phosphatases at concentrations as low as 20-25 nM, induces apoptosis of R- mouse embryo fibroblasts, which are 3T3-like cells devoid of type 1 insulin-like growth factor receptors (IGF-IRs). From R- cells, we have generated (by stable transfection) cell lines with IGF-IR numbers ranging from 0 (R- cells) to >10(6) receptors per cell. The wild-type IGF-IR protects R- cells from OKA-induced apoptosis, its protective effect being exquisitely dependent on the number of receptors. A small increment in wild-type receptor number (from 15 x 10(3) to 22 x 10(3) receptors/cell) is sufficient to change R(-)-derived cells from sensitive to resistant to apoptosis. We have also studied the effect of various mutations of the IGF-IR on its ability to protect R(-)-derived cells from OKA-induced apoptosis. Our data indicate a correlation between protection from apoptosis and the ability of the receptor to respond to insulin-like growth factor I with mitogenesis.

The role of Grb2 in the growth and transformation of mouse embryo cells.

An overexpressed insulin-like growth factor I receptor (IGF-IR) allows cells to grow in IGF-I only and to form colonies in soft agar. Conversely, cells with a targeted disruption of the IGF-IR genes, R- cells, are refractory to transformation by several oncoproteins and growth factor receptors, that readily transform their wild type counterparts, W cells. Grb2 is an SH2-SH3 domains protein that links tyrosine kinase receptors to ras signalling. In order to determine its role in mitogenesis and transformation, we have transfected a plasmid expressing Grb2 into R- and W cells, and their derivatives already expressing the SV40 large T antigen. In addition, we have used loss-of-function mutants of Grb2 to inquire whether they would act as dominant negatives. Our results show that: (1) an overexpressed Grb2 cannot replace the IGF-IR in IGF-I-mediated mitogenesis; (2) Grb2 also fails to transform either W or R- cells; (3) Grb2 and SV40 T antigen, singly transfected, cannot transform R- cells, but can do so when combined; and (4) SH3 domain mutants of Grb2 act as dominant negatives, causing reversion of the transformed phenotype. We conclude that Grb2 is necessary but not sufficient for transformation.

Mutational analysis of the mitogenic and transforming activities of the insulin-like growth factor I receptor.

THe type 1 insulin-like growth factor receptor (IGF-IR) plays an important role in mitogenesis and transformation. It has been previously shown that mitogenic signaling and transforming activity of the IGF-IR can be dissociated: a receptor truncated at residue 1229 (C-terminus) is fully mitogenic, in terms of its response to IGF-I, but cannot transform 3T3-like cells that are devoid of endogenous IGF-IRs (R- cells). We have extended our mutational analysis of the C-terminus of the human IGF-IR, by stably transfecting several mutant receptors into R- cells, and testing the resulting cell lines for IGF-I-mediated mitogenic response and formation of colonies in soft agar. The results indicate that the transforming domain of the IGF-IR can be localized between residues 1245 and 1310, these sequences being not required for mitogenic signaling. Within these residues, there are at least two areas that contribute to the transforming activity of the receptor.

A differential proteomic approach to identify proteins associated with thyroid cell transformation.

Tumour suppressor p53 is a transcription factor essential for DNA damage checkpoints during cellular response to stress. Mutations in the p53 gene are the most common genetic alterations found in human tumours; most pathogenetic modifications are missense mutations that abolish the p53 DNA-binding function. In the same cell type, distinct p53 missense mutations may determine different phenotypes. The PC Cl3 cell line retains several markers of thyroid differentiation in vitro. Introduction of the V143A mutant p53 allele, which abolishes the p53 DNA-binding function, leads to loss of differentiation markers as well as TSH dependency for growth. Conversely, PC Cl3 cells transfected with the S392A mutant p53 allele, presenting the mutation located outside the DNA-binding domain, show only loss of TSH dependency for growth. To identify molecular differences existing between PC Cl3 cell lines transformed by the V143A and the S392A mutant alleles, a differential proteomic approach was used. Two-dimensional gel electrophoresis analyses indicated that expression of a significant portion of protein species was modified by both p53 mutants. In fact, compared with wild-type PC Cl3 cells, modification of expression in V143A mutant cells occurred in 23.6% of the entire protein species. Conversely, modification of S392A mutant cells affected 14.0% of total proteins. Among these components, 8.3% were common to both mutants. Several of these proteins were identified by mass spectrometry procedures; some proteins, such as HSP90 and T-complex proteins, are already known to be related to p53 function.

Inhibition of tumor growth by a dominant negative mutant of the insulin-like growth factor I receptor with a bystander effect.

The insulin-like growth factor I receptor is known to play a major role in transformation and apoptosis. The dominant negative mutant of the insulin-like growth factor I receptor, designated 486/STOP, causes massive apoptosis of tumor cells and inhibition of tumor growth and metastases. We now show that: (a) the stable expression of 486/STOP inhibits transformation (colony formation in soft agar) and/or tumor growth in nude mice of five different types of human tumor cell lines; and (b) more importantly, it has a bystander effect, inhibiting the growth of wild-type tumor cells when cells expressing 486/STOP are coinjected with wild-type tumor cells. These findings suggest that it is not necessary to infect all tumor cells with 486/STOP to inhibit tumor growth, and they also open the possibility of using the product of 486/STOP directly against tumor cells.

Multiple plasma proteins control atrial natriuretic peptide (ANP) aggregation.

We have recently demonstrated that human alpha-atrial natriuretic peptide (alpha-hANP), an amyloidogenic peptide responsible for isolated atrial amyloidosis, binds to a dimeric form of apo A-I belonging to small high-density lipoproteins (HDL). This binding phenomenon is considered a protective mechanism since it inhibits or strongly reduces the ANP aggregation process. The observation that plasma exhibits at least four times greater amyloid inhibitory activity than HDL prompted us to determine whether small HDL are the only ANP plasma-binding factors. After incubation of whole plasma with labelled ANP, the macromolecular complexes were subjected to two-dimensional gel electrophoresis followed by autoradiography. The results presented here provide novel evidence of additional binding proteins, in addition to apo A-I dimer, able to bind ANP in vitro and to prevent its aggregation. The mass spectrometry analysis of the radioactive spots identified them as albumin, alpha-1 antitrypsin, orosomucoid and apo A-IV-TTR complex. The putative impact of these findings in the amyloidogenic/antiamyloidogenic peptides network is discussed.

In vitro reactivity of penicilloyl and penicillanyl albumin and polylysine conjugates with IgE-antibody.

Penicilloylated (BPO) and penicillanylated (BPA) poly-L-lysine (PLL) and human serum albumin (HSA) were prepared and characterized by penamaldate assay and proton NMR spectroscopy. The conjugates were coupled to nitrocellulose (NC) discs and cyanogen bromide activated paper discs and their in vitro reactivities with serum IgE antibodies were examined. Results showed that on paper discs, 55.3 and 83% of the sera reacted with PLL conjugates of BPO and BPA, respectively, while 41.5 and 58.1% reacted with HSA conjugates. On NC discs, HSA conjugates gave better results, 75.6 and 70.7%, respectively for BPO and BPA, compared with 38.6 and 50%, respectively for the PLL conjugates. Overall, the BPA-PLL conjugate on paper discs proved to be the most reactive preparation. Addition of the BPO-PLL paper disc preparation detected more positive sera (85.1%) and we believe that the combined use of these two specificities offers the best test for the detection of penicillin-reactive IgE antibodies.

Sleep-related breathing disorders and cardiovascular disease.

Sleep-related breathing disorders, ranging from habitual snoring to the increased upper airway resistance syndrome to sleep apnea, are now recognized as major health problems. The majority of patients have excessive daytime sleepiness and tiredness. Neuropsychological dysfunction results in poor work performance, memory impairment, and even depression. Until recently, the coexistence of cardiovascular and cerebrovascular diseases with sleep-related breathing disorders was thought to be the result of shared risk factors, such as age, sex, and obesity. However, in the past 5 years several epidemiologic studies have demonstrated that sleep-related breathing disorders are an independent risk factor for hypertension, probably resulting from a combination of intermittent hypoxia and hypercapnia, arousals, increased sympathetic tone, and altered baroreflex control during sleep. Sleep apnea may lead to the development of cardiomyopathy and pulmonary hypertension. Early recognition and treatment of sleep-related breathing disorders may improve cardiovascular function.

Investigation of conformational specificity at GPIIb/IIIa: evaluation of conformationally constrained RGD peptides.

RGD-containing proteins and peptides are known to bind to the platelet GPIIb/IIIa receptor and inhibit platelet aggregation. That a conformational component to the specificity exists is suggested by significantly lower activity of linear RGD analogs relative to closely related cyclic peptides and small proteins containing the RGD sequence. Recently, conformations for a suite of RGD containing cyclic peptides have been defined by NMR-based methods and, for one molecule, by X-ray diffraction. We report here the NMR-based conformational analysis of an additional cyclic peptide, cyclo(Pro-Arg-Gly-Asp-D-Pro-Gly), and compare the conformational variations in the suite of peptides and related analogs. Biological activity data for these peptides shows a preference of the platelet GPIIb/IIIa receptor for one conformation of the RGD sequence, but suggests its ability to bind a second, distinct conformation.

High plasma levels of endothelin-1 in untreated Addison's disease.

The aim of this study has been to investigate the plasma endothelin-1 (ET-1) levels in adult patients with proven Addison's disease (AD). Plasma ET-1 levels were measured in 29 subjects (17 males and 12 females, aged between 20 and 54 years): 15 of them were patients with AD and 14 were sex- and age-matched normal subjects, used as a control group. All patients with AD have been studied under basal conditions and nine of them also after 2 weeks on oral corticosteroid therapy (individual cortisol dosage ranging from 25 to 37.5 mg/day and 0.1 mg/day 9 alpha-fluorohydrocortisone). Extracted plasma ET-1 was determined by a specific radioimmunoassay using rabbit endothelin antisera. Mean ET-1 values in the patients with AD were three times higher than in normal subjects (21.09 +/- 4.38 pg/ml vs 6.72 +/- 1.74 pg/ml; p < 0.0001). Plasma ET-1 levels assayed in the patients with AD after 2 weeks of corticosteroid therapy were significantly decreased (14.47 +/- 3.7 pg/ml vs 22.8 +/- 5.2 pg/ml; -37%; p < 0.001) compared to values in untreated patients. However, the plasma ET-1 values obtained following corticosteroid therapy were still significantly higher (p < 0.001) than those detected in the control subjects. These results clearly indicate that patients with untreated AD have increased circulating ET-1 levels that may be reduced by short-term corticosteroid therapy.

Quality of life in patients with obstructive sleep apnea: effect of nasal continuous positive airway pressure--a prospective study.

BACKGROUND: Obstructive sleep apnea (OSA) is a common condition and is associated with excessive daytime sleepiness and neuropsychological dysfunction. There is limited evidence on the effect of OSA on the quality of life and its response to nasal continuous positive airway pressure (nCPAP) treatment. STUDY OBJECTIVE: To determine the effect of nCPAP on the quality of life in patients with OSA. DESIGN: Prospective determination of nCPAP effect in a case-series analysis. PATIENTS: We studied 29 patients (23 were male and 6 were female) with a mean (+/-SE) age of 4.4+/-2.3 years, a body mass index 36.3+/-2.0 kg/height (m)2, and a diagnosis of OSA with respiratory disturbance index (RDI; apnea/hypopnea) of 77+/-9 events/h. MEASUREMENTS AND RESULTS: The quality of life was assessed by administering a Medical Outcomes Study Short Form-36 questionnaire before and after 8 weeks of nCPAP therapy in polysomnographically documented OSA. All dimensions of the quality of life were significantly impaired when compared with an age- and gender-matched population, expressed as a percentage of normative data: physical functioning, 75%; vitality, 41%; role functioning (physical, 54%; emotional, 61%; social, 66%); general health, 88%; and mental health, 76%. nCPAP therapy significantly improved the sleep-disordered breathing and sleep fragmentation. The nCPAP level for the group was 9.4+/-0.7 cm H2O. Eight weeks of nCPAP therapy improved vitality (75%), social functioning (90%), and mental health (96%). The magnitude of improvement was related to the degree of quality of life impairment prior to treatment, rather than to the severity of disease as measured by the RDI and arousal indices. CONCLUSIONS: All aspects of the quality of life, from physical and emotional health to social functioning, are markedly impaired by OSA. nCPAP therapy improved those aspects related to vitality, social functioning, and mental health.

Increased plasma levels of endothelin-1 in patients with hyperthyroidism.

Plasma concentrations of endothelin-1 (ET-1) were measured in 25 hyperthyroid subjects, 15 hypothyroid subjects, and 21 age-matched normal controls. In hyperthyroid patients, plasma concentrations of ET-1 were significantly higher than in the control group (P < .0001) and in hypothyroid patients (P < .0001). In contrast, no differences were found between hypothyroid patients and controls. Plasma levels of ET-1 were similarly elevated as in patients with Graves' disease and those with toxic adenoma. No correlations were found between plasma ET-1 levels, thyroid hormones, and thyrotropin (TSH) in hyperthyroid, hypothyroid, and euthyroid groups. The results of our study clearly indicate that in hyperthyroidism, circulating levels of ET-1 are strongly increased, although the pathogenesis of the increase is unclear.