Hard tissue lesion treatment in oral and maxillofacial has been challenging because of tissue complexities. This study aimed to investigate novel biopolymeric construct effects on the osteogenic differentiation potential of the dental pulp stem cells (DPSCs) for introducing a cell copolymer bioimplant. A blended polycaprolactone (PCL)‐polyethylene oxide (PEO) was fabricated using electrospinning, simultaneously filled by β‐glycerophosphate (β‐GP). After that biocompatibility and release kinetics of the PCL‐PEO+β‐GP was evaluated and compared with PCL‐PEO and then the osteogenic differentiation potential of the DPSCs was examined while being cultured on the scaffolds and compared with those cultured on the culture plate. The results demonstrated that scaffolds have not any cytotoxicity and β‐GP can release in a long‐term manner. Alkaline phosphatase activity and calcium content were significantly increased in DPSCs while being cultured on the PCL‐PEO+β‐GP compared with the other groups. Runt‐related transcription factor 2, collagen type‐I, osteonectin, and osteocalcin (OSC) genes expression was upregulated in DPSCs cultured on the PCL‐PEO+β‐GP and was significantly higher than those cultured on the PCL‐PEO. Immunocytochemistry result also confirmed the positive effects of PCL‐PEO+β‐GP on the osteogenic differentiation of the DPSCs by presenting a higher OSC protein expression. According to the results, incorporation of the β‐GP in PCL‐PEO makes a better construct for osteogenic induction into the stem cells and it could be also considered as a great promising candidate for bone, oral, and maxillofacial tissue engineering applications.

Skin is the most frequently affected organ in acute graft versus host disease (GVHD). Data from murine studies support the hypothesis that the interaction of residing host Langerhans cells of the epidermis (LC) and donor T cells is crucial for the initiation of acute GVHD. Donor T cells are also necessary to induce the switch of LC from host to donor origin after allogeneic stem cell transplantation (SCT). In an ongoing clinical protocol applying alemtuzumab-based T cell depleted (TCD) allogeneic SCT (Meyer, Blood2007; 109:374), we observed acute skin GVHD occurring early after transplantation. We therefore intended to analyse the LC chimerism in patients undergoing this protocol. So far, LC-chimerism analysis in humans has been performed by the detection of the Y-chromosome restricting it to sex-mismatched donor/recipient pairs. Here we introduce a new method to isolate LC from small skin samples at high purity for a sensitive STR-based chimerism analysis of general applicability. Epidermal skin layers were obtained from 6 mm punch biopsies by dispase I digestion. A small slice of epidermis was used for immunofluorescent staining. The remaining sample was digested by trypsin, and CD1a/MHC-class II-positive LC were sorted by flow cytometry. This approach resulted in a mean purity of > 96% with skin of healthy individuals. However, the density of LC early after SCT following non-TCD myeloablative regimens had previously been shown to be much lower compared to healthy individuals. By CD1a-staining, we were able to show that this is also the case after TCD reduced intensity SCT. Nevertheless, LC could be purified in all of 8 analyzed patients. The isolated LC numbers ranged from 10 to more than 1000. In 4 patients we performed a re-analysis of the isolated cells by flow cytometry and confirmed a purity exceeding 97%. We obtained reliable results for LC chimerism in 6 of 8 patients. After the RNA-isolation protocol was further improved, we were able to detect signals even with 35 isolated LC in patient MZ-47. In two patients, the majority of isolated LC were of donor origin whereas the other 4 patients had predominantly host LC (patients’ characteristics and chimerism results are summarized in Table 1). None of the patients developed spontaneous acute GVHD so far. For patients MZ-37 and MZ-43 LC-chimerism was also performed after day +50 post HSCT and showed a switch to >97% donor chimerism at that time. In summary, we have established a sensitive method that enables the chimerism analysis on highly purified LC independent of sex-mismatched donor/recipient pairs. Our results on a few patients’ samples can not yet be related to clinical events. This assay, however, allows the comprehensive investigation of the chimerism of LC and potentially of other tissue-resident antigen presenting cells to study their impact on GVHD in humans. Table 1 patient donortype sex (patient/donor) LC (n) purity (%) donor chimerism (%) MSD, matched sibling donor; MMUD, mismatched unrelated donor; n.a., not applicable; M, male; F, female 37 MSD F/M 1127 97,7 <3 39 MMUD F/M 750 98,6 >97 40 MMUD M/M 1800 98,3 13 43 MMUD M/M 10 n.a. n.a. 44 MSD M/M 157 n.a. n.a. 45 MSD M/F 325 > 98 36 46 MMUD M/M 355 n.a. 48 47 MSD M/M 35 n.a. >90