Histological analysis of bone formation and revascularization in the osteotomy gap.

<p>(<b>A</b>) Photomicrographs of histological sections 6 weeks post-surgery of the CD133(+) (left) and the ABC (right) group (Movat Pentachrome staining); In the CD133(+) group the osteotomy gap was predominantly filled with spongy mineralized tissue formations. A thin layer of hyaline cartilage (green) between the bone ends had not yet been mineralized. In the ABC group almost only fibrous tissue was found, completely lacking any cartilage or bone formation; (ABC: autologous blood clot group; CD133(+): CD133 cell treated group); cortical bone/yellow; bone marrow/dark red; cartilage/green; muscle/red <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0052650#pone.0052650-BPreininger1" target="_blank">[21]</a>. (<b>B</b>) Photomicrographs of immune histological sections 6 weeks post-surgery of the CD133(+) (left) and the ABC (right) group (factor VIII staining); In the CD133(+) group quite dense vessel formations (red) were observed, especially in the endostal region at the zone where the hyaline cartilage formations are mineralized. (ABC: autologous clot group; CD133: CD133+ cell treated group) (Scale bar: 1 mm).</p

µCT analysis – 3D reconstructions of the bone healing progress.

<p>(<b>A</b>) 3D reconstructions from µCT scans from one representative animal after 2, 4, and 6 weeks of the three groups are shown. In the CD133(+) group (left) mineralized tissue was formed at the endosteal as well as at the periosteal regions. After 42 days the gap had almost been bridged; a thin layer of radiolucent tissue still separated the bone ends. In the ABC group (middle) only very little callus formation was observed, appearing between the 28th and 42nd day after the osteotomy. No bridging but a cap-formation was observed, sealing the medullary canal. In the CD34(+) group (right) bone formation was further progressed than in the ABC group. After 6 weeks however, bridging did not occur and the medullary canal was closed at the proximal end. (Resolution 28 µm; scale: 2 mm). (<b>B</b>) Bridging score for the ABC group is lower compared with the bridging in the CD34(+) or CD133(+) group. The bridging score for the CD34(+) group showed better results than the ABC group but lesser results compared with the CD133(+). 5 areas in the osteotomy gap were investigated, 12, 3, 6, and 9 o'clock position and the centre region. The sum of bridged areas for the healing course is indicated in the table.</p

<i>In vitro</i> analysis of angiogenic potential of progenitor cells.

<p>(A) Tube length evaluation for CD34 (+)/(−) and CD133(+)/(−) cells normalized to the tube length measured in the control group together with all PBMCs, note the significant difference in tube length between CD133(+) and CD133(−), n = 8, * p = 0.018; (B) Tube formation of CD133(+) and CD133(−) respectively, note the more pronounced tube formation in the CD133(+) fraction.</p

µCT analysis – statistic evaluation of callus volume and bone mineral contend.

<p>An increase of callus volume (CV, [mm3]), as well as bone mineral contend (BMC, [mg]) on the basis of total mineral density of the callus was observed over time. Both parameters were significantly higher in the CD133(+) group when compared to the ABC group. Note that the differences occur not until after the 28th day and are most pronounced after the 42nd day. Comparing CV and BMC between the CD133(+) group and the CD34(+) group markedly higher values were found in the CD133(+) group. (CV: total volume, BMC: bone mineral content, wks: weeks); (CD133(+) group  =  grey; CD34(+) group  =  checkered; ABC (autologous blood clot) group  =  white).</p

data_sheet_1_Distinct Housing Conditions Reveal a Major Impact of Adaptive Immunity on the Course of Obesity-Induced Type 2 Diabetes.docx

<p>Obesity is associated with adipose tissue inflammation, insulin resistance, and the development of type 2 diabetes (T2D). However, our knowledge is mostly based on conventional murine models and promising preclinical studies rarely translated into successful therapies. There is a growing awareness of the limitations of studies in laboratory mice, housed in abnormally hygienic specific pathogen-free (SPF) conditions, as relevant aspects of the human immune system remain unappreciated. Here, we assessed the impact of housing conditions on adaptive immunity and metabolic disease processes during high-fat diet (HFD). We therefore compared diet-induced obesity in SPF mice with those housed in non-SPF, so-called “antigen exposed” (AE) conditions. Surprisingly, AE mice fed a HFD maintained increased insulin levels to compensate for insulin resistance, which was reflected in islet hyperplasia and improved glucose tolerance compared to SPF mice. By contrast, we observed higher proportions of effector/memory T cell subsets in blood and liver of HFD AE mice accompanied by the development of non-alcoholic steatohepatitis-like liver pathology. Thus, our data demonstrate the impact of housing conditions on metabolic alterations. Studies in AE mice, in which physiological microbial exposure was restored, could provide a tool for revealing therapeutic targets for immune-based interventions for T2D patients.</p