Influence of body condition score and its change after parturition on pregnancy rates to fixed-timed artificial insemination in Bos indicus beef cows.

This experiment evaluated pregnancy rates to fixed-time artificial insemination (FTAI) in Bos indicus beef cows according to their body condition score (BCS) at calving and subsequent change until 30 days after FTAI. Non-pregnant, suckling Nelore cows (n = 593 primiparous, 461 secundiparous, and 893 multiparous) were evaluated for BCS at calving and FTAI, and at 30 days after FTAI when cow pregnancy status was verified. Cow BCS at calving was subtracted from BCS recorded at pregnancy diagnosis, and cows classified as those that lost BCS (L), maintained BCS (M), or gained BCS (G) during this period. Cows that calved with BCS ≥ 5.0 had greater (P < 0.01) BCS throughout the experiment, and greater (P < 0.01) pregnancy rates to FTAI compared with cows that calved with BCS < 5.0 (54.8 vs. 34.2%). Pregnancy rates to FTAI were greater (P < 0.01) for G and M cows compared with L cows (50.0%, 47.5%, and 36.0%, respectively), and similar (P = 0.46) between G and M cows. Moreover, pregnancy rates to FTAI in G cows that calved with BCS < 5.0 were less compared with L (tendency; P = 0.08) and M cows (P < 0.01) that calved with BCS ≥ 5.0 (42.2%, 48.3%, and 58.3%, respectively). In summary, pregnancy rates to FTAI were greater in B. indicus cows that calved with a BCS ≥ 5.0 regardless of parity and post-calving BCS change, and greater in M and G cows within those that calved with BCS < 5.0 or ≥ 5.0.

Pelvic organ prolapse repair-relapse risk factors: A 10-year retrospective study / Reparación del prolapso de órganos pélvicos: factores de riesgo de recaída. Estudio retrospectivo de 10 años

ObjectivesPelvic organ prolapse (POP) is a common problem among older female. The usual treatment for POP is surgery but there are high recurrence rates, with a 29% reoperation rate. This study aims to identify risk factors for both primary prolapse and recurrence after surgical treatment.MethodsRetrospectively assessment of clinical records of patients who underwent surgery for POP in a 10-year period. Statistical analysis was performed using the version 26.0 of Statistical Package for the Social Science (SPSS®) software.Results746 women entered our study. The population was predominantly post-menopausal, multiparous, and obese/overweight. The most affected compartment was the anterior. Almost 90% of the patient presented with major prolapse. Being overweight or obese, having apical compartment POP, major POP or all compartment POP were risk factors for recurrence with statistical significance. The recurrence rate was nearly one-third but the reoperation incidence was low, reaching less than 6%.ConclusionsPOP surgery has a high satisfaction rate. The only modifiable risk factor for recurrence is being overweight/obese and a nutritional plan should be considered before surgery so we can achieve the best possible results.

ObjetivosEl prolapso de órganos pélvicos (POP) es un problema común entre las mujeres. El tratamiento habitual del POP es la cirugía, pero existen altas tasas de recurrencia, con una tasa de reintervención del 29%. Este estudio tiene como objetivo identificar los factores de riesgo tanto para el prolapso primario como para la recurrencia después del tratamiento.MétodosEvaluación retrospectiva de historias clínicas de pacientes intervenidos por POP en un período de 10 años. El análisis estadístico se realizó utilizando la versión 26.0 del software Statistical Package for the Social Science (SPSS®).ResultadosIngresaron a nuestro estudio 746 mujeres. La población era predominantemente posmenopáusica, multípara y obesa/con sobrepeso. El compartimento más afectado fue el anterior. Casi el 90% de los pacientes presentó prolapso mayor. Tener sobrepeso u obesidad, tener POP de compartimento apical, POP mayor o POP de todos los compartimentos fueron factores de riesgo de recurrencia con significación estadística. La tasa de recurrencia fue de casi un tercio, pero la incidencia de reintervención fue baja, alcanzando menos del 6%.ConclusionesLa cirugía POP tiene un alto índice de satisfacción. El único factor de riesgo modificable de recurrencia es el sobrepeso/obesidad y se debe considerar un plan nutricional antes de la cirugía para que podamos lograr los mejores resultados posibles.

BIO-NAIL: a bacterial cellulose dressing as a new alternative to preserve the nail bed after avulsion.

Avulsion of the nail plate is the most accomplished surgical procedure among the nail apparatus surgeries. Since it is not possible to use the removed nail to cover the nail bed, some materials have been suggested, however, they are generally not available for use and often at a high cost. To evaluate the use of Bacterial Cellulose (BC) dressing as a biological nail (Bio-Nail) after partial or total avulsion of the nail plate. Twenty-six candidates for nail avulsion, were randomized into two groups: Control, using Vaseline with gauze (11 patients) and Experimental group, BC group, using the Bio-Nail (15 patients). The patients were followed up during the 180-day period. The distribution of the patients in the groups was homogeneous for both sociodemographic and clinical data. The occurrence of infection (1 case in the control group) was not statistically relevant. The BC group had lower pain intensity (p = 0.011) with earlier temporal resolution when compared to the control group (p = 0.003). The BC group presented earlier reepithelization (p = 0.022) and better quantitative (p = 0.021) and qualitative conditions (p = 0.011) for the exudate. Regarding satisfaction, all the patients were satisfied. Good preservation of the nail plate area was observed in the BC group at the end of the 180-day period (p = 0.024). Average time of BC dressing permanence was 16.4 ± 7.1 days. BC showed to be appropriate as a dressing after partial or total avulsion of the nail plate. BC is a Bio-Nail promising for nail bed healing.

Galectin-3 orchestrates the histology of mesentery and protects liver during lupus-like syndrome induced by pristane.

Galectin-3 (Gal-3) controls intercellular and cell-extracellular matrix interactions during immunological responses. In chronic inflammation, Gal-3 is associated with fibrotic events, regulates B cell differentiation and delays lupus progression. Gal-3 deficient mice (Lgals3-/-) have intense germinal center formation and atypical plasma cell generation correlated to high levels IgG, IgE, and IgA. Here, we used pristane (2,6,10,14-tetramethylpentadecane) to induce lupus-like syndrome in Lgals3-/- and Lgals3+/+ BALB/c mice. Mesentery and peritoneal cells were monitored because promptly react to pristane injected in the peritoneal cavity. For the first time, mesenteric tissues have been associated to the pathogenesis of experimental lupus-like syndrome. In Lgals3+/+ pristane-induced mice, mesentery was hallmarked by intense fibrogranulomatous reaction restricted to submesothelial regions and organized niches containing macrophages and B lymphocytes and plasma cells. In contrast, Lgals3-/- pristane-treated mice had diffuse mesenteric fibrosis affecting submesothelium and peripheral tissues, atypical M1/M2 macrophage polarization and significant DLL1+ cells expansion, suggesting possible involvement of Notch/Delta pathways in the disease. Early inflammatory reaction to pristane was characterized by significant disturbances on monocyte recruitment, macrophage differentiation and dendritic cell (DC) responses in the peritoneal cavity of pristane-induced Lgals3-/- mice. A correlative analysis showed that mesenteric damages in the absence of Gal-3 were directly associated with severe portal inflammation and hepatitis. In conclusion, it has suggested that Gal-3 orchestrates histological organization in the mesentery and prevents lupoid hepatitis in experimental lupus-like syndrome by controlling macrophage polarization, Notch signaling pathways and DC differentiation in mesenteric structures.

Efeito da suplementação parenteral extra de cobre e zinco sobre a resposta imunológica de vacas Nelore / Effects of parenteral over-supplementation of cooper and zinc on immune response of Nellore cows

O objetivo deste estudo foi avaliar o efeito da suplementação mineral injetável extra de cobre (Cu) e zinco (Zn) sobre a resposta imunológica de vacas Nelore no período pré-parto. Foram avaliadas 60 vacas prenhes, as quais foram divididas em dois tratamentos, por meio da distribuição aleatória em delineamento inteiramente ao acaso. Aos 75 dias antes do parto, as vacas do tratamento testemunha (T) receberam soro fisiológico como placebo e os animais suplementados (S) receberam mineral injetável via subcutânea (75mg de cobre e 250mg de zinco, em dose única). Foram realizadas três coletas de sangue, duas antes da data prevista para o parto (75 e 10 dias) e uma 30 dias após o parto. Os teores de Cu, Zn, ceruloplasmina, imunoglobulinas G (IgG) e M (IgM) foram analisados durante as três coletas. A atividade fagocitária foi avaliada aos 30 dias pós-parto. Os dados foram examinados mediante análise de variância, com o uso do pacote estatístico do SAS, e os dados individuais da atividade fagocitária pelo PROC GLM. Os dados de Cu, Zn, IgG e IgM foram analisados como medidas repetidas no tempo de coleta por meio do PROC MIXED, com o nível de significância de 5%. Os teores de Cu, Zn, IgM, IgG, ceruloplasmina e a atividade fagocitária das vacas não sofreram influência dos tratamentos (P>0,05). O fornecimento de Cu e Zn injetável, nas doses utilizadas, aos 75 dias antes do parto para vacas Nelore, em dietas suficientes, não alterou os teores de Cu, Zn, ceruloplasmina e a resposta imunológica até 30 dias após o parto.(AU)

The aim of this study was to evaluate the effect of extra injectable mineral supplementation of copper (Cu) and zinc (Zn) on the immune response of Nellore cows in pre-partum period. Sixty pregnant cows were randomly distributed in a completely randomized design in two treatments. In the control treatment (T), cows received saline as placebo, and supplemented treatment (S) received mineral injection (75mg copper and 250mg of zinc, single dose) subcutaneously, 75 days prior to parturition. Blood was sampled three times, two before the expected date of parturition (75 and 10 days) and another at 30 days postpartum. Analyses were performed for Cu, Zn, ceruloplasmin, immunoglobulin G (IgG) and M (IgM) in the three periods and the phagocytic activity in the last period (30 days postpartum). The experimental data were subjected to analysis of variance using the statistical package SAS, being that the individual data phagocytic activity were analyzed by PROC GLM, and the Cu, Zn, IgG and IgM were analyzed as repeated measures in the time, using the PROC MIXED, with the significance level of 5%. The Cu, Zn, IgM, IgG, ceruloplasmin and the phagocytic activity of the cows were not affected by treatments (P>0.05). The supply of injectable Cu and Zn, at the doses used, 75 days before parturition to Nellore cows in sufficient diets, did not alter the serum contents of Cu, Zn, ceruloplasmin and the immune response up to 30 days after parturition.(AU)

Effects of temperament on physiological, productive, and reproductive responses in beef cows.

This experiment evaluated the effects of temperament on physiological, productive, and reproductive responses in beef cows. A total of 953 lactating, multiparous, non-pregnant Nelore cows (age = 99 ± 2 mo; days post-partum = 51.4 ± 0.3 d; BCS = 5.34 ± 0.04; BW = 430 ± 2 kg) were allocated into 8 groups of approximately 120 cows each. Groups were assigned to an estrus synchronization + timed-AI protocol at the beginning of the breeding season. Concurrently with AI, blood samples were collected, hair samples were clipped from the tail switch, and cow temperament was evaluated via chute score and exit velocity. Individual exit score was calculated within each group by dividing exit velocity into quintiles and assigning cows with a score from 1 to 5 (1 = slowest; 5 = fastest cow). Temperament scores were calculated by averaging cow chute score and exit score, and used to define cow temperament ( ≤ 3 = adequate, = 726; ADQ; > 3 = excitable, = 227; EXC). Cows not pregnant to AI were assigned to a second timed-AI protocol ( = 184 ADQ and 72 EXC) or exposed ( = 269 ADQ and 90 EXC) to bulls for 60 d. Pregnancy status was verified 30 d after each AI and 45 d after the breeding season via transrectal ultrasound. Cow age, BW, BCS, and d post-partum at the beginning of the breeding season were similar ( ≥ 0.27) between ADQ and EXC cows. At first timed-AI, EXC had greater ( < 0.01) serum cortisol but similar ( ≥ 0.87) serum haptoglobin and hair cortisol concentrations compared with ADQ cows (49.1 vs. 39.1 ng/mL of serum cortisol, SEM = 1.0). Pregnancy rate to first timed-AI tended ( = 0.09) to be less in EXC vs. ADQ cows (41.0 vs. 47.3%; SEM = 3.6), whereas no treatment differences were detected ( ≥ 0.23) for subsequent pregnancy outcomes. Calving rate was less ( = 0.04) in EXC vs. ADQ cows (68.3 vs. 74.8%; SEM = 2.2), which can be attributed to the greater ( = 0.05) pregnancy loss detected in EXC cows (9.9 vs. 5.9%; SEM = 1.4). Weaning rate tended ( = 0.09) to be less, whereas calf weaning BW and age were less ( ≤ 0.05) in EXC vs. ADQ cows (63.9 vs. 69.4%, SEM = 2.4; 209 vs. 212 d, SEM = 1; 204 vs. 210 kg, SEM = 2). Hence, kg of calf weaned/cow exposed to breeding was reduced ( = 0.04) in EXC vs. ADQ cows (130 vs. 146 kg, SEM = 5). In summary, cows with excitable temperament had reduced reproductive performance and overall productivity compared to cohorts with adequate temperament when exposed to timed-AI + natural breeding.

Impacts of meloxicam administration before temporary calf weaning on physiological and reproductive responses of beef cows.

The objective of this experiment was to evaluate temperament, physiological, and reproductive variables in beef cows assigned to an estrus synchronization + timed AI protocol including eCG administration, 48-h temporary calf weaning (TCW), or TCW + meloxicam administration. A total of 943 lactating, multiparous, nonpregnant Nelore cows, allocated into 8 groups of approximately 120 cows each, were assigned to the experiment. Groups were maintained in individual pastures and assigned to the following estrus synchronization + timed AI protocol: a 2-mg injection of estradiol benzoate and an intravaginal progesterone-releasing device (CIDR) on d 0, a 12.5-mg injection of PGF on d 7, CIDR removal in addition to a 0.6-mg injection of estradiol cypionate on d 9, and timed AI on d 11. Within each group, cows were randomly assigned on d 9 to 1) TCW from d 9 to 11 (TCW-CON; = 317), 2) no TCW and a 300-IU injection of eCG on d 9 (NOTCW; = 311), and 3) TCW-CON in addition to meloxicam administration (intramuscular; 0.5 mg/kg BW) on d 9 (TCW-MEL; = 315). Cow BW and BCS were assessed on d 0. On d 9 and 11, blood samples were collected, and cow temperament was evaluated via chute score and exit velocity. Pregnancy status was verified 30 d after timed AI via transrectal ultrasonography. No treatment differences were detected ( ≥ 0.23) for cow age, days postpartum, BW, and BCS on d 0 of the estrus synchronization + timed AI protocol. No treatment effects were detected ( ≥ 0.41) for any of the temperament variables evaluated. A treatment × day interaction was detected ( = 0.02) for serum cortisol concentrations, which were similar ( = 0.55) between treatments on d 9 but greater ( ≤ 0.05) in TCW-CON and TCW-MEL compared with NOTCW cows on d 11. No treatment effects were detected ( = 0.90) for serum haptoglobin concentrations, which decreased from d 9 to 11 in all treatments (day effect; < 0.01). No treatment differences were detected ( = 0.84) for pregnancy rates to timed AI. In summary, TCW during estrus synchronization did not impact temperament or serum haptoglobin concentrations in beef cows but increased serum cortisol concentrations compared with cows not assigned to TCW, although such an outcome was not sufficient to impact pregnancy rates to timed AI. Moreover, administration of meloxicam did not alleviate the TCW-induced increase in serum cortisol concentrations and failed to benefit pregnancy rates to timed AI in beef cows.

Neuromyelitis optica spectrum disorders: beyond longitudinally extensive transverse myelitis.

AIM: To describe the neuroradiological features and their prevalence in patients with neuromyelitis optica (NMO). MATERIALS AND METHODS: Two neuroradiologists independently reviewed 35 spinal cord and 37 brain MRI studies from patients with NMO. The examinations were analysed for the presence of lesion, topography, enhancement, and brain lesions suggestive of multiple sclerosis and/or NMO. RESULTS: Seventy percent of the spinal cord lesions involved over three or more vertebral segments. Seventy-eight percent of brain scans were abnormal, and the most prevalent findings were non-specific foci of T2 hyperintensities in the cerebral white matter (55%) and brainstem lesions (52%). One patient had lesions disseminated in space compatible with multiple sclerosis according to 2010 revised McDonald criteria. Brain lesions suggestive of NMO occurred at least once in 17 (59%) patients. CONCLUSION: Spinal cord lesions were often longitudinally extensive and brain lesions were common, with the majority of patients having at least one distinctive NMO lesion.

Efficacy of dental floss impregnated with chlorhexidine on reduction of supragingival biofilm: a randomized controlled trial.

BACKGROUND: The use of a toothbrush has a limited ability to control the dental biofilm in interproximal areas. Therefore, specialized devices, such as dental floss, may be useful for these specific areas. OBJECTIVE: This study aimed to investigate the efficacy of dental floss impregnated with 5% chlorhexidine gluconate on the reduction of the supragingival biofilm. METHODS: This research was parallel, single-blind, controlled and randomized, and contained a sample of thirty dental students from the Faculty of Pharmacy, Dentistry and Nursing of the Federal University of Ceará, Brazil, who were divided equally into three groups. The negative control group (NC) did not utilize any kind of interproximal cleaning; the positive control group (PC) used waxed floss without impregnation twice a day; and the test group (T) used the same dental floss, which was impregnated with 5% chlorhexidine gluconate, twice a day. For all groups, this study lasted for 15 days. The presence of a biofilm was evaluated on four surfaces (mesiobuccal, distobuccal, mesiolingual and distolingual) by the Quigley-Hein Index, resulting in four scores for each tooth. RESULTS: Group T had the lowest plaque scores, showing a significant difference compared to group NC (P < 0.001) and group PC (P < 0.001). Group PC also displayed a significant difference compared to NC (P < 0.001). CONCLUSION: It was concluded that the use of dental floss impregnated with 5% chlorhexidine gluconate resulted in additional reductions in the supragingival biofilm relative to the results achieved with conventional waxed floss on the anterior teeth of a well-motivated and well-instructed population.

Evaluation of light-touch sensation in the buccal mucosa of leprosy patients.

OBJECTIVE: The aim of this study is to evaluate the light-touch sensation of the oral mucosa in leprosy patients. MATERIALS AND METHODS: A cross-sectional study included 228 adults, 133 being leprosy patients and 95 normal controls. To assess light-touch sensation, the five-filament standard Semmes-Weinstein kit in eight regions of the oral mucosa was used. Chi-square test was used to verify the differences in responses between the studied groups. RESULTS: Normal sensation was predominant in both groups, and diminished sensation was found also in the nonleprosy group. Normal controls showed diminished light-touch sensation in at least one point, which may be attributed to the definition of sensation normality. The chi-square test revealed no significant difference between the two groups (p = 0.43171). Severe diminished sensation was detected only in the leprosy group. CONCLUSION: It is concluded that altered light-touch sensation in the oral cavity may not be a common feature in leprosy. CLINICAL RELEVANCE: Decreased or lost sensation in skin lesions is a paramount in leprosy diagnosis; however, clinicians must be aware that this seems not to be true in the oral mucosa in leprosy cases for diagnosis purpose.

Evaluation of light-touch sensation in the buccal mucosa of leprosy patients

OBJECTIVE: The aim of this study is to evaluate the light-touch sensation of the oral mucosa in leprosy patients. MATERIALS AND METHODS: A cross-sectional study included 228 adults, 133 being leprosy patients and 95 normal controls. To assess light-touch sensation, the five-filament standard Semmes-Weinstein kit in eight regions of the oral mucosa was used. Chi-square test was used to verify the differences in responses between the studied groups. RESULTS: Normal sensation was predominant in both groups, and diminished sensation was found also in the nonleprosy group. Normal controls showed diminished light-touch sensation in at least one point, which may be attributed to the definition of sensation normality. The chi-square test revealed no significant difference between the two groups (p = 0.43171). Severe diminished sensation was detected only in the leprosy group. CONCLUSION: It is concluded that altered light-touch sensation in the oral cavity may not be a common feature in leprosy.CLINICAL RELEVANCE: Decreased or lost sensation in skin lesions is a paramount in leprosy diagnosis; however, clinicians must be aware that this seems not to be true in the oral mucosa in leprosy cases for diagnosis purpose.

Genetic diversity of polysporic isolates of Moniliophthora perniciosa (Tricholomataceae).

The causal agent of witches' broom disease, Moniliophthora perniciosa is a hemibiotrophic and endemic fungus of the Amazon basin and the most important cocoa disease in Brazil. The purpose of this study was to analyze the genetic diversity of polysporic isolates of M. perniciosa to evaluate the adaptation of the pathogen from different Brazilian regions and its association with different hosts. Polysporic isolates obtained previously in potato dextrose agar cultures of M. perniciosa from different Brazilian states and different hosts (Theobroma cacao, Solanum cernuum, S. paniculatum, S. lycocarpum, Solanum sp, and others) were analyzed by somatic compatibility grouping where the mycelium interactions were distinguished after 4-8 weeks of confrontation between the different isolates of M. perniciosa based on the precipitation line in the transition zone and by protein electrophoresis through SDS-PAGE. The diversity of polysporic isolates of M. perniciosa was grouped according to geographic proximity and respective hosts. The great genetic diversity of M. perniciosa strains from different Brazilian states and hosts favored adaptation in unusual environments and dissemination at long distances generating new biotypes.

The role of angiogenesis in a murine tibial model of distraction osteogenesis.

Distraction osteogenesis (DO) is one of the most dramatic in vivo applications of mechanical stimulation as a means of inducing bone regeneration. A simple and reproducible murine model of tibia distraction osteogenesis was developed using a monolateral fixator. Bone formation was assessed histologically over a 35-day time course. The steady state expression of a broad family of angiogenesis-associated genes was assessed by microarray hybridization analyses over the same time course, while the immediate gene response that was induced during each cycle of distraction was assessed at 30 min and 8 h after the first and last rounds of activation of the fixator. Distraction osteogenesis promoted new bone formation primarily through an intramembranous process with maximal osteogenesis during the active distraction period. Histological analysis also showed that dense cortical bone continued to be formed, during the consolidation phase, for 2 weeks after distraction ended. The analysis of steady state mRNA expression levels over the time course of DO showed that VEGF-A and neuropilin, an alternate receptor for VEGF-A, both angiopoietin (Ang) 1 and 2 factors, and the Ang receptor Tie2 were the critical angiogenic factors during DO. A key transcriptional regulator of many of the angiogenic factors, hypoxia-induced factor1alpha (Hif-1a), the FGF binding protein pleiotropin/OSF1, and multiple MMP(s), were also induced during the active distraction period. Examination of the expression of angiogenic factors that were induced after each cycle of activation, demonstrated that Hif-1a, Nrp1, and VEGF-A were all cyclically induced after each increment of distraction. These results suggest that these factors are early mediators that are produced by distraction and contribute toward the processes that promote bone formation. These experiments represent the first step in defining the molecular mechanisms that regulate skeletal regeneration and the functional relationship between angiogenesis and osteogenesis during distraction osteogenesis.

Selective adhesion of osteoblastic cells to different integrin ligands induces osteopontin gene expression.

Skeletal homeostasis is partly regulated by the mechanical environment and specific signals generated by a cell's adhesion to the matrix. Previous studies demonstrated that osteopontin (OPN) expression is stimulated in response to both cellular adhesion and mechanical stimulation. The present studies examine if specific integrin ligands mediate osteoblast selective adhesion and whether opn mRNA expression is induced in response to these same ligands. Embryonic chicken calvaria osteoblastic cells were plated on bacteriological dishes coated with fibronectin (FN), collagen type I (Col1), denatured collagen/gelatin (G), OPN, vitronectin (VN), laminin (LN) or albumin (BSA). Osteoblastic cells were shown to selectively adhere to FN, Col1, G and LN, yet not to VN, OPN or BSA. Opn mRNA expression was induced by adhesion to Col1, FN, LN and G, but neither OPN nor VN induced this expression. Examination of the activation of the protein kinases A and C second signaling systems showed that only adhesion to FN induced protein kinase A and protein kinase C (PKC) activity while adherence to Col1 induced PKC. Evaluation of the intracellular distribution of focal adhesion kinase (FAK) and p-tyrosine within cells after adherence to FN, VN or BSA demonstrated that adherence to FN stimulated FAK translocation from the nucleus to the cytoplasm and high levels of p-tyrosine localization at the cell surface. However, cell adherence to VN or BSA did not show these morphological changes. These data illustrate that osteoblast selective adhesion is mediated by specific integrin ligands, and induction of intracellular second signal kinase activity is related to the nature of the ligand.

Predominant integrin ligands expressed by osteoblasts show preferential regulation in response to both cell adhesion and mechanical perturbation.

Previous studies have demonstrated that both mechanical perturbation and cell adhesion induced the expression of osteopontin (opn) by osteoblasts (Carvalho et al. [1998] J. Cell. Biochem. 70:376-390). The present study examined if these same stimuli on osteoblasts would induce the expression of other integrin binding proteins, specifically fibronectin (fn) and bone sialoprotein (bsp). All three genes showed three- to four-fold maximal induction in response to both cell adhesion and a single 2-h period of an applied spatially uniform, dynamic biaxial strain of 1.3% at 0.25 Hz. Each gene, however, responded with a different time course of induction to mechanical strain, with bsp, fn, and opn showing their maximal response at 1, 3, and 9 h, respectively, after the perturbation period. In contrast, peak induction to cell adhesion was observed at 24 h for bsp and opn, while fn levels peaked at 8 h. Interestingly, while both opn and fn mRNA expression returned to base line after cell adhesion, bsp mRNA levels remained elevated. Examination of collagen type I and osteocalcin mRNAs showed unaltered levels of expression in response to either type of perturbation. A common feature of the signal transduction pathways, which mediate the gene expression in response to both cell adhesion and mechanical perturbation, was the activation of specific tyrosine kinases based on the ablation of the induction of these genes by the tyrosine kinase inhibitor genistein. While cycloheximide blocked the induction of all three mRNAs in response cell adhesion, it failed to block the induction of any of these genes in response to mechanical perturbation. Such results suggest that the induction of these genes after mechanical perturbation was mediated by an immediate response to signal transduction, while cell adhesion mediated effects secondary to signal transduction. Depolymerization of microfilaments with cytochalasin D had no effect on the overall expression of any of these genes in response to cell adhesion and only blocked the induction of opn expression in response to mechanical perturbation. These results suggest that cytoskeletal integrity is only selectively important in the signal transduction of certain types of stimuli and for the regulation of certain genes. In summary, both mechanical perturbation and cell adhesion stimulated the expression of integrin binding proteins. Furthermore, while there are common features in the signal transduction processes that mediate the induction of these genes in response to both stimuli, specific genes are separately regulated by precise mechanisms that are unique to both forms of stimuli.

Osteoblasts induce osteopontin expression in response to attachment on fibronectin: demonstration of a common role for integrin receptors in the signal transduction processes of cell attachment and mechanical stimulation.

Osteopontin is a predominant integrin binding protein of bone and its expression has been shown to be induced by mechanical stimuli within osteoblasts (Toma et al. [1997] J. Bone Miner. Res. 12:1626-1636). The present studies examined if the cell adhesion would mimic the mechano-transduction that stimulated opn mRNA expression and whether integrin receptors were involved in these processes. Osteopontin mRNA expression was induced three- to four-fold, 24 hours after embryonic chicken calvaria osteoblast attachment to fibronectin (FN), however no induction was observed if the cells were plated on tissue culture plastic alone. Osteopontin mRNA induction in response to cell attachment on FN was dependent on new protein synthesis and the activation of a tyrosine protein kinase(s) but unlike mechano-induction was independent of the maintenance of the cell's microfilament structure. Integrin receptor(s) were shown to be involved in mediating the signal transduction processes of both cell attachment and mechanical stimulation since incubation of osteoblasts with the integrin binding peptide RGDS partially blocked the induction of opn expression in response to both stimuli. Interestingly, incubation of the osteoblasts that were adherent on tissue culture plastic alone with the RGDS peptide lead to an induction in opn expression suggesting that integrin occupancy by itself was sufficient to initiate the signal transduction process that induced opn expression. In order to assess the role of integrin occupancy vs. focal adhesion complex formation that accompanies cell attachment, in the signal transduction process that induces opn expression, receptor clustering was stimulated pharmacologically with bombesin or lysophosphatidic acid in osteoblasts attached to tissue culture plastic. Neither compound in the absence of occupancy of the integrin receptors was capable of stimulating opn expression in attached cells, however if the cells were placed in suspension pharmacological mediation of receptor clustering and integrin occupancy were additive in their effect of inducing opn expression. These data demonstrate that induction of opn expression by mechanical stimuli and cell attachment are commonly mediated through integrin receptor(s). However, when cells are attached receptor clustering alone which accompanies focal adhesion formation was incapable of mediating signal transduction suggesting that receptor occupancy was a prerequisite to the signal transduction process that leads to the induction of opn mRNA expression.

Collagen synthesis from human PDL cells following orthodontic tooth movement.

The dynamic remodelling processes in the periodontal ligament (PDL) account for the reaction of PDL cells to different orthodontic force simulations. These occur mostly by degradation and synthesis of collagen types I, III, V, VI, XII and XIV. The purpose of this study was to quantify specific collagen types in the PDL from zones of tension and compression of experimental teeth. Such changes could then be correlated with the processes of orthodontic-stimulated tissue breakdown. Maxillary and mandibular premolars of three females and one male patient were orthodontically moved with a box loop for a total of 14 days, prior to tooth extraction. Teeth from the contralateral side of either the maxilla or the mandible served as the untreated controls. A total of seven experimental and seven control teeth were used in this investigation. PDL fibroblasts from the cervical third of the roots corresponding to the compression and tension zones of the experimental and control teeth, respectively, were scraped and cultured in vitro at 37 degrees C in a humidified incubator with 5 per cent CO2/95 per cent air. Collagen synthesis of types I, III, V and VI was quantified by using an ELISA. Application of orthodontic forces in the experimental teeth showed a significant increase (P < 0.05) of the synthesis of all collagen types in the compression as opposed to the tension zones. Collagen synthesis on the compression zone of experimental teeth was not significantly different in the mandible when compared with those of the maxilla. In addition, the proportional distribution of different types of collagen was also not significantly different in the PDL fibroblasts from either zone of experimental teeth of either the maxilla or the mandible. Collagen metabolism in response to orthodontic stimulation appears to be higher in the compression zones and lower in the tension zones. Contrary to what is traditionally assumed in the literature, such findings indicate that in addition to bone resorption, tissue remodelling is very active in zones of compression following the disappearance of the hyalinized areas. These findings constitute a model for future studies on collagen metabolism during orthodontic-stimulated tooth movement.

Conformational changes upon binding of a receptor loop to lipid structures: possible role in signal transduction.

The mas oncogene codes for a seven transmembrane helix protein. The amino acid sequence 253-266, from the third extracellular loop and beginning of helix 7, was synthesized either blocked or carrying an amino acid spin label at the N-terminus. Peptide binding to bilayers and micelles was monitored by ESR, fluorescence and circular dichroism. Binding induced tighter lipid packing, and caused an increase of peptide secondary structure. While binding to bilayers occurred only when peptide and phospholipid bore opposite charges, in micelles the interaction took place irrespective of charge. The results suggest that changes in lipid packing could modulate conformational changes in receptor loops related to the triggering of signal transduction.

Glycosaminoglycan synthesis in the rat articular disk in response to mechanical stress.

The mechanism by which compressive mechanical stress affects glycosaminoglycan synthesis in the articular disk was investigated with a modified organ culture technique. Forty-eight male Sprague-Dawley rats were divided into three experimental groups and one control group of 12 animals each, aged 7 and 9 weeks. The experimental groups followed different regimens of stress applied for 25%, 75%, or 100% of the time during the total test period of 24 hours. Articular disks were stressed with flexible bottomed dishes (Flex I dishes, Flexcell Corp., McKeesport, Pa.) using the Flexercell Strain Unit (Flexcell Corp., McKeesport, Pa.) and incubated with [3H]-glucosamine for 24 hours. Samples were then collected, digested with Pronase-E, and after precipitation with cetylpyridinium chloride (CPC) and ethanol, the different glycosaminoglycans (GAGs) were separated by using cellulose acetate electrophoresis. The significant GAG types with stress were chondroitin6sulfate (C6S), hyaluronic acid (HA), and dermatan sulfate (DS). There was no significant relationship in the experimental groups between age and regimen of stress applied in either age. Higher stress regimens showed significantly higher proportions of C6S when compared with the controls, whereas HA appeared to decrease slightly and DS was not affected. Since C6S is the major component of hyaline cartilage, the results of this study suggest that compressive forces in the articular disk may stimulate the development of more cartilagenous-like properties with respect to GAG content.