Diabetes is the major risk factor for nontraumatic lower extremity amputation (LEA). The role of genetic polymorphisms in predisposing diabetics to impaired wound healing leading to LEA has not been sufficiently explored. We investigated the association between a set of genes belonging to the angiogenesis/wound repair pathway with LEA in the Chronic Renal Insufficiency Cohort, a study of adults with chronic kidney disease (CKD) that includes a subgroup with diabetes. This study was performed on 3,772 Chronic Renal Insufficiency Cohort participants who were genotyped on the ITMAT-Broad-CARe array chip. A total of 1,017 single-nucleotide polymorphisms (SNPs) in 22 genes belonging to the angiogenesis/would repair pathway were investigated. LEA was determined from patient self-report. The association between genetic variants and LEA status was examined using logistic regression and additive genetic models after stratifying the cohort by race/ethnicity and diabetic status. Unadjusted analyses as well as analyses adjusted for age, sex, estimated glomerular filtration rate, body mass index, peripheral vascular disease, hemoglobin A1c, and population stratification were performed. In non-Hispanic white participants with diabetes, rs11938826 and rs1960669, both intronic SNPs in the gene basic fibroblast growth factor-2 (FGF2), were significantly associated with LEA in covariate-adjusted analysis (OR: 2.83 (95% CI: 1.73, 4.62); p-value: 0.000034; Bonferroni adjusted p-value: 0.0006) and (OR: 2.61 (95% CI: 1.48, 4.61); p-value: 0.00095; Bonferroni adjusted p-value: 0.02). In the same subgroup, rs10883688, an FGF8 SNP of unknown functional effect, was also associated with LEA (OR: 1.72 (95% Confidence Interval: 1.14, 2.6); p-value: 0.00999; Bonferroni adjusted p-value: 0.04). No statistically significant associations were identified in the other ethnic groups. In conclusion, variant/s in FGF2 and FGF8 may predispose diabetics with CKD to LEA. Dysregulation of the FGF2 gene represents an opportunity to understand further, and possibly intervene upon, mechanisms of wound healing in diabetics with CKD.

OBJECTIVE: The aim of this study was to differentiate human embryonic stem cells (hESCs) into odontoblastic lineage in an optimized culture milieu.
METHODS: In Phase 1, hESCs were differentiated into mesenchymal stem cells (H9-MSCs). In Phase 2, H9-MSCs were then differentiated into odontoblast-like cells (H9-Odont) under the stimulation of FGF-8 and BMP-4. Alternatively, H9-MSCs were differentiated into osteogenic lineage (H9-Osteo). In Phase 3, H9-Odont were seeded on 17% EDTA-treated dentine substrates in the presence of FGF-8 and BMP-4 for further differentiation. All experiments were performed in triplicate (n = 3). One-way anova was used to test hESC differentiation into different cell types. Post hoc Tukey\'s test was used to compare between groups. P < 0.05 was considered statistically significant.
RESULTS: H9-Odont expressed the odontoblastic marker DSPP gene 125.47 ± 0.1 (SD)-folds higher compared with H9-MSCs at mRNA level (real-time RT-PCR). Additionally, the flow cytometry results revealed 53.1 ± 3.4 (SD) % of DSP (+) cells in H9-Odont. Alternatively, H9-Osteo expressed 5.9 ± 2.2 (SD) % of DSP (+) cells. Moreover, the SEM results demonstrated that H9-Odont were found to undergo morphological changes from a fibroblast-like shape into more rounded shapes with cytoplasmic extensions into the dentinal tubules when seeded on 17% EDTA-treated dentine substrate in the presence of FGF-8 and BMP-4. However, H9-Osteo and H9-MSCs did not show similar morphological changes under similar culture milieu.
CONCLUSIONS: This study supports the potential of hESCs as a stable, consistent, unlimited and \'off-the-shelf\' cell source to obtain odontoblastic cells for future clinical and research applications.

OBJECTIVE: To evaluate cellular hypertrophic activities in the mandibular condylar cartilage (MCC) and the glenoid fossa (GF) during mandibular advancement in the temporomandibular joint (TMJ) of Sprague-Dawley rats, as evidenced by fibroblast growth factor 8 (FGF8).
METHODS: Fifty-five female 24-day-old Sprague-Dawley rats were randomly divided into four experimental and control groups, with a mandibular advancement appliance on the experimental rats\' lower incisors. The rats were euthanized on days 3, 14, 21, and 30 of the study, and their TMJ was prepared for a immunohistochemical staining procedure to detect FGF8.
RESULTS: FGF8 expression was significantly higher among the experimental rats (P  =  .002). Patterns of ascension and descension of FGF8 expression were similar in experimental and control samples. The results show an overall enhanced osteogenic transition occurring in both the MCC and the GF in experimental rats in comparison with controls. The level of cellular changes in the MCC is remarkably higher than in the GF.
CONCLUSIONS: In the MCC and the GF, cellular morphologic and hypertrophic differentiations increase significantly during mandibular advancement. It is also concluded that endochondral ossification in the MCC and intramembranous ossification in the GF occur during adaptive remodeling.

OBJECTIVE: To investigate the effects of FGF-8 on cranial neural crest cell (CNCC) differentiating into ectomesenchymal cell of the first branchial arch, and determine the appropriate dose and stage of CNCC exposure to FGF-8.
METHODS: Cranial neural crest explants were cultured in free-serum medium containing modified DMEM/F12 and different doses of FGF-8. The differentiation type of CNCC were determined by in situ hybridization for Hoxa2 and immunocytochemistry for vimentin.
RESULTS: Pre-emigrating CNCC demonstrated the negative Hoxa2 stain and positive vimentin stain after treated by 100 ug/FGF-8. Both post-emigrating CNCC group and control group were positive for Hoxa2 and vimentin stain.
CONCLUSIONS: On the early stage of CNCC emigration, the first branchial arch phenotype of CNCC could be induced by FGF-8. This experiment could provide in vitro model for study on the mechanism of tooth-jaw regeneration.

Mitogen-activated protein kinase (MAPK) pathways are well known to be involved in signal transduction from extracellular to intracellular compartments in all eukaryotes. The activation of this cascade will have an effect on cell proliferation, differentiation, and apoptosis. In this study, we describe the cloning of the chick Mkp3 gene that is highly homologous to the mammalian gene and are both expressed in several embryo regions with demonstrated morphogenetic activity. In early developmental stages, Mkp3 and Fgf8 have similar expression patterns. Differences in the activation of Mkp3 transcription in the isthmus and the repression with FGF receptor inhibition suggest that Fgf8 protein controls Mkp3 transcription. Ectopically, expression of Fgf8 protein induces Mkp3 in a short period of time in the diencephalon, indicating a positive regulation of Mkp3 by Fgf8. Moreover, we show a distinct tissue competence to express Mkp3 rostrally and caudally to the zona limitans intrathalamica (ZLI).

Fibroblast growth factor 15 (Fgf15) is a gene regulated by the expression of Otx2 in developing mouse brain (Proc Natl Acad Sci USA 97 (2000) 14388). Otx2 gene codes for a transcription factor and is fundamental for the regionalisation and development of the anterior neural plate and cephalic region of the vertebrate embryo (Development 124 (1997) 3639). In addition, the thalamic expression of Fgf15 has been recently reported under the control of Shh signalling gene, expressed in the diencephalic basal plate (Development 129 (2002) 4807). In the present work, we have analysed Fgf15 expression pattern during mouse neural development. Fgf15 appeared early in the developing neural epithelium, in domains where Fgf8 gene is also expressed and, at later stages, in specific groups of neural cells. Fgf8 is an important signalling protein with demonstrated morphogenetic activity in several embryonic regions. Fgf15 expression is localized, like Fgf8, in secondary neural tube organizers: the isthmic organizer (IsO) and the anterior neural ridge (ANR).