Microbiome-induced Increases and Decreases in Bone Tissue Strength can be Initiated After Skeletal Maturity.

Recent studies in mice have indicated that the gut microbiome can regulate bone tissue strength. However, prior work involved modifications to the gut microbiome in growing animals and it is unclear if the same changes in the microbiome, applied later in life, would change matrix strength. Here we changed the composition of the gut microbiome before and/or after skeletal maturity (16 weeks of age) using oral antibiotics (ampicillin + neomycin). Male and female mice (n=143 total, n=12-17/group/sex) were allocated into five study groups:1) Unaltered, 2) Continuous (dosing 4-24 weeks of age), 3) Delayed (dosing only 16-24 weeks of age), 4) Initial (dosing 4-16 weeks of age, suspended at 16 weeks), and 5) Reconstituted (dosing from 4-16 weeks following by fecal microbiota transplant from Unaltered donors). Animals were euthanized at 24 weeks of age. In males, bone matrix strength in the femur was 25-35% less than expected from geometry in mice from the Continuous (p= 0.001), Delayed (p= 0.005), and Initial (p=0.040) groups as compared to Unaltered. Reconstitution of the gut microbiota, however, led to a bone matrix strength similar to Unaltered animals (p=0.929). In females, microbiome-induced changes in bone matrix strength followed the same trend as males but were not significantly different, demonstrating sex-related differences in the response of bone matrix to the gut microbiota. Minor differences in chemical composition of bone matrix were observed (Raman spectroscopy). Our findings indicate that microbiome-induced impairment of bone matrix in males can be initiated and/or reversed after skeletal maturity. The portion of the femoral cortical bone formed after skeletal maturity (16 weeks) is small; however, this suggests that microbiome-induced changes in bone matrix occur without osteoblast/osteoclast turnover using an, as of yet unidentified mechanism. These findings add to evidence that the mechanical properties of bone matrix can be altered in the adult skeleton.

In Vitro Assessment of Human Islet Vulnerability to Instant Blood-Mediated Inflammatory Reaction (IBMIR) and Its Use to Demonstrate a Beneficial Effect of Tissue Culture.

Culture of human pancreatic islets is now routinely carried out prior to clinical islet allotransplantation, using conditions that have been developed empirically. One of the major causes of early islet destruction after transplantation is the process termed instant blood-mediated inflammatory reaction (IBMIR). The aim of this study was to develop in vitro methods to investigate IBMIR and apply them to the culture conditions used routinely in our human islet isolation laboratory. Freshly isolated or precultured (24 h, 48 h) human islets were incubated in either ABO-compatible allogeneic human blood or Hank's buffered salt solution (HBSS) for 1 h at 37°C. Tissue factor (TF) expression and leukocyte migration were assessed by light microscopy. TF was also quantified by ELISA. To assess ß-cell function, glucose-stimulated insulin secretion (GSIS) assay was carried out. The extent of islet ß-cell damage was quantified using a proinsulin assay. Islets cultured for 24 h had higher GSIS when compared to freshly isolated or 48-h precultured islets. Freshly isolated islets had significantly higher TF content than 24-h and 48-h precultured islets. Incubation of freshly isolated human islets in allogeneic human blood released 6.5-fold higher level of proinsulin in comparison to freshly isolated human islets in HBSS. The high level of proinsulin released was significantly attenuated when precultured islets (24 h or 48 h) were exposed to fresh blood. Histological examination of fresh islets in blood clot showed that some islets were fragmented, showing signs of extraislet insulin leakage and extensive neutrophil infiltration and necrosis. These features were markedly reduced when the islets were cultured for 24 h. These results suggest that our standard 24-h islet culture is markedly beneficial in attenuating IBMIR, as evidenced by increased GSIS, lower content of TF, decrease islet fragmentation, and proinsulin release.

Functional recovery ten years after pediatric traumatic brain injury: outcomes and predictors.

Functional impairments (adaptive, behavioral, educational) are common after preschool traumatic brain injury (TBI). In comparison with cognitive outcome, functional outcomes have received limited attention, with little evidence to determine whether these difficulties persist in the long term. The aim of this study was to examine functional outcomes at 10 years post-injury and identify predictors of outcome. The study compared children with a diagnosis of TBI (n=40) to a healthy age-, gender-, and socioeconomic status (SES)-matched control group (n=19) at 10 years post-injury. Outcomes and predictors of functional skills were investigated. Poorer adaptive skills were evident for those with more severe injury. Behavioral difficulties were present regardless of injury severity. Post-injury, arithmetic skills were the most compromised in the longer term. Pre-injury status, interventions accessed, and acute intellectual function were significant predictors of outcome. These results highlight the importance of monitoring functional skills in the long term, especially for those children presenting with risk factors.

The biomechanical characteristics of the bone-periodontal ligament-cementum complex.

The relative motion between the tooth and alveolar bone is facilitated by the soft-hard tissue interfaces which include periodontal ligament-bone (PDL-bone) and periodontal ligament-cementum (PDL-cementum). The soft-hard tissue interfaces are responsible for attachment and are critical to the overall biomechanical efficiency of the bone-tooth complex. In this study, the PDL-bone and PDL-cementum attachment sites in human molars were investigated to identify the structural orientation and integration of the PDL with bone and cementum. These attachment sites were characterized from a combined materials and mechanics perspective and were related to macro-scale function. High resolution complimentary imaging techniques including atomic force microscopy, scanning electron microscopy and micro-scale X-ray computed tomography (Micro XCT) illustrated two distinct orientations of PDL; circumferential-PDL (cir-PDL) and radial-PDL (rad-PDL). Within the PDL-space, the primary orientation of the ligament was radial (rad-PDL) as is well known. Interestingly, circumferential orientation of PDL continuous with rad-PDL was observed adjacent to alveolar bone and cementum. The integration of the cir-PDL was identified by 1-2 microm diameter PDL-inserts or Sharpey's fibers in alveolar bone and cementum. Chemically and biochemically the cir-PDL adjacent to bone and cementum was identified by relatively higher carbon and lower calcium including the localization of small leucine rich proteins responsible for maintaining soft-hard tissue cohesion, stiffness and hygroscopic nature of PDL-bone and PDL-cementum attachment sites. The combined structural and chemical properties provided graded stiffness characteristics of PDL-bone (E(r) range for PDL: 10-50 MPa; bone: 0.2-9.6 GPa) and PDL-cementum (E(r) range for cementum: 1.1-8.3 GPa), which was related to the macro-scale function of the bone-tooth complex.

Structure of the integrin binding fragment from fibrillin-1 gives new insights into microfibril organization.

Human fibrillin-1, the major structural protein of extracellular matrix (ECM) 10-12 nm microfibrils, is dominated by 43 calcium binding epidermal growth factor-like (cbEGF) and 7 transforming growth factor beta binding protein-like (TB) domains. Crystal structures reveal the integrin binding cbEGF22-TB4-cbEGF23 fragment of human fibrillin-1 to be a Ca(2+)-rigidified tetragonal pyramid. We suggest that other cbEGF-TB pairs within the fibrillins may adopt a similar orientation to cbEGF22-TB4. In addition, we have located a flexible RGD integrin binding loop within TB4. Modeling, cell attachment and spreading assays, immunocytochemistry, and surface plasmon resonance indicate that cbEGF22 bound to TB4 is a requirement for integrin activation and provide insight into the molecular basis of the fibrillin-1 interaction with alphaVbeta3. In light of our data, we propose a novel model for the assembly of the fibrillin microfibril and a mechanism to explain its extensibility.

Changing patterns of mitral stenosis in childhood and pregnancy in Sri Lanka.

The findings in rheumatic mitral stenosis appear to have undergone changes, probably in association with improved socioeconomic conditions, in developing countries. The objective of this study was to assess such changes and to adapt strategies of management. The clinical and pathologic features, mortality rate, long-term functional class and restenosis rate in 168 children and 62 pregnant women who underwent closed transventricular mitral valvotomy in the first 14-year period (June 1964 to May 1978) were compared and correlated with those of 140 children and 106 pregnant women in the following 7-year period (June 1978 to May 1985). During the late period, there were attenuated severity of the disease, emergence of a mild pathologic type of valve involvement confined to the commissures (commissural band stenosis), decreased mortality (1.2%) and restenosis rates (p less than 0.001) and increased long-term improvement (p less than 0.001) during childhood. The best results of closed valvotomy were obtained in simple commissural and commissural band stenosis, the latter forming the predominant group in children and pregnant women in the late period (p less than 0.001). Poor results were observed in patients with the subtype of combined stenosis characterized by commissural rigidity, cuspal stenosis and chordal fusion, demonstrating the inapplicability of closed valvotomy. Closed valvotomy is safe in all stages of pregnancy, as evidenced by the zero mortality rate and rate (1.8%) of fetal death, and offers good long-term palliation. However, in pregnant women with pure mitral stenosis characterized by simple commissural or commissural band stenosis, balloon valvuloplasty is an acceptable alternative, especially in light of the risks associated with surgery. The choice of the procedure for the relief of stenosis is determined by the pathologic anatomy of the valve stenosis.

An evaluation of factors relevant to the diagnosis of bronchial carcinoma in Sri Lanka.

The incidence of bronchial carcinoma in Sri Lanka is low. The extent to which its pattern varies from that prevailing in western countries is demonstrated by a study of 84 cases diagnosed over a 10-year period. Of the patients examined 45-2 per cent were non-smokers, all 17 women belonging to this category. A negative smoking history, especially in women, was not a useful guide in assessing a suspected lesion. Adenocarcinoma, which is usually peripheral, comprised a high proportion of the tumours (22-6%). The majority of cases (70-3%) had had peripheral tumours which were beyond bronchoscopic visibility. Therefore, bronchoscopy was not as useful a diagnostic procedure as in the West. Digital clubbing was seen in less than half the males, while only two females had this sign.

Some observations of oesophageal carcinoma in Ceylon, including its relationship to betel chewing.

A series of 237 cases of oesophageal carcinoma admitted to two thoracic units in Ceylon is analysed.Evidence suggestive of an aetiological link between betel chewing and high incidence of the tumour in Ceylon is presented. The sex incidence is unusual in that there is a preponderance of females in the series. A significant proportion of patients were women under 40 years of age. The middle third of the oesophagus was the commonest site affected.