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Optimal in vitro conditions are necessary for the development of a strong, well structured, and functional tissue engineered cardiovascular structure eventually designed for implantation. To further optimize in vitro conditions for cell proliferation and extracellular matrix formation in tissue engineering of cardiovascular structures, in this study, ascorbic acid and growth factors as additives to standard cell culture medium were evaluated for their effect on tissue development in vitro. Biodegradable polymer patches [polyglycolic acid (PGA) coated with poly-4-hydroxybutyrate (P4HB)] were seeded with human pediatric aortic cells and cultured for 7 and 28 days. Group A was cultured with standard medium (DMEM with 10% fetal calf serum and 1% antibiotics) supplemented with ascorbic acid; group B was cultured with standard medium plus ascorbic acid and basic fibroblast growth factor (bFGF); group C was cultured with standard medium adding ascorbic acid and transforming growth factor (TGF). Analysis of the cell seeded polymer constructs included DNA assay, collagen assay, and histologic and immunohistochemical examination for cell proliferation and collagen formation. After 7 and 28 days of culture, group B and group C showed a significantly higher DNA content compared with group A. The addition of bFGF (group B) led to a markedly higher collagen synthesis after 28 days of culture compared with the additives in groups C and A. The histologic and immunohistochemical examination also revealed a more dense, organized tissue development with pronounced matrix protein formation in the tissue engineered structures in group B after 28 days of culture. When seeded on to the polymeric scaffold, human vascular cells proliferate and form organized cell tissue after 28 days of culture. The addition of bFGF and ascorbic acid to the standard medium enhances cell proliferation and collagen synthesis on the biodegradable polymer, which leads to the formation of more mature, well organized tissue engineered structures.

Summary Background Striate palmoplantar keratoderma (SPPK; OMIM #148700) is a rare autosomal dominant genodermatosis characterized by linear hyperkeratosis on the digits and hyperkeratosis on the palms and soles. SPPK is known to be caused by heterozygous mutations in either the desmoglein 1 (DSG1), desmoplakin (DSP), or keratin 1 (KRT1) genes. Objective To define the molecular basis of SPPK in five Pakistani families showing a clear autosomal dominant inheritance pattern of SPPK. Methods Based on previous reports of DSG1 mutations in SPPK, we performed direct sequencing of the DSG1 gene of all five families. Results Mutation analysis resulted in the identification of one recurrent mutation (p.R26X) and four novel mutations (c.Ivs4-2A>G, c.515C>T, c.Ivs9-3C>G, and c.1399delA) in the DSG1 gene. Each mutation is predicted to cause haploinsufficiency of DSG1 protein. Conclusion The results of our study further underscore the significance of the desmoglein gene family in diseases of epidermal integrity.

Abstract(1) Background-Atrichia with papular lesions (APL) is a rare autosomal recessive form of inherited alopecia. Affected individuals present with a distinct pattern of total hair loss on the scalp, axilla and body shortly after birth and are essentially devoid of eyelashes and eyebrows. This form of hair loss is irreversible and the histology is consistent with an absence of mature hair follicles. In addition to total atrichia, APL patients also present with papules and follicular cysts filled with cornified material. Mutations in the Hairless (HR) gene have been shown to underlie APL.(2) Objective-Here, we studied five unrelated large Pakistani families with clinical manifestations of APL.(3) Methods-Based on previous reports of HR mutations in APL, we performed direct DNA sequencing analysis.(4) Results-DNA sequencing of the HR gene in APL patients revealed three novel nonsense mutations in five unrelated families. All affected individuals were homozygous for a nonsense mutation due to C-to-T transitions at different positions in the amino acid sequence. Two families carry the mutation Q323X (CAG-TAG) in exon 3, two families harbor the mutation Q502X (CAG-TAG) in exon 6, and one family had a mutation at R940X (CGA-TGA) in exon 14. Haplotype analysis revealed that all affected individuals of both APL1 and APL16 families were homozygous for the same haplotype, and likewise, the mutation in families APL2 and APL19 was on the the same haplotype.(5) Conclusions-We report three novel nonsense mutations in the HR gene in APL. Two of the newly identified mutations, Q323X and Q502X, were found to be shared between unrelated families and marker analysis confirmed an identical homozygous haplotype for APL1 and APL16, and for APL2 and APL19. These findings suggest that Q323X and Q502X did not arise independently, but instead appear to have been propagated in the population. Collectively, these findings contribute further evidence for the involvement of hairless mutations in papular atrichia.

Background-Keratins are heteropolymeric proteins that form the intermediate filament cytoskeleton in epithelial cells. The common basic structure of all keratins is organized in a central α-helical rod domain flanked by nonhelical, variable head and tail regions. Most mutations in keratins are found in the central α-helical rod domain. Keratin 9 (K9) is expressed only in the suprabasal layers of palmoplantar epidermis. Mutations in the keratin 9 gene (KRT9) have been shown to cause epidermolytic palmoplantar keratoderma (EPPK; OMIM 144200), an autosomal dominant genodermatosis characterized clinically by diffuse hyperkeratosis limited to palms and soles and histologically by epidermolysis in suprabasal layers of the epidermis.