In situ-RT-PCR and immunohistochemistry for the localisation of the mRNA of the alpha 3 chain of laminin and laminin-5 during human organogenesis

Abstract

Laminin-5 is known to be an integral part of the hemidesmosome and therefore responsible for the integrity of the connection of the epithelium to the basement membrane. This is also an important mechanism during embryonic development, as documented by studies in mice. In an attempt to elucidate its implication for human development we localised the mRNA of the alpha3 chain of laminin with the help of in situ RT-PCR, and the laminin-5 protein immunohistochemically. We systematically investigated kidney, lung, skin and intestinal tissue of consecutive developmental stages during human embryogenesis. From gw 6.5 onwards, the mRNA of the alpha3 chain of laminin was found exclusively in the cytoplasm of epithelial cells of the developing kidney, lung, skin and intestine. Interestingly, in the skin and intestine from gw 8 onwards, the superficial cell layers also stained positive for the mRNA, while the protein was still only found in the dermal-epidermal and enteric basement membrane zones. In all developing organs investigated, the mRNA of the alpha3 chain of laminin is strictly of epithelial origin and the corresponding protein localised in the underlying basement membrane zones. Due to this discrepancy, we postulate a broader role for laminin-5 during human embryogenesis, for example, for epithelial cell development, beyond its involvement in hemidesmosome formation and cell adhesion.

Identification of a Novel Family of Laminin N-terminal Alternate Splice Isoforms

“…Negligible signal was obtained in connective tissues. These results largely parallel those previously obtained for LAMA3 transcripts in skin and hair (39, 40). …”

Adhesion and Migration, the Diverse Functions of the Laminin α3 Subunit

“…Placenta expresses the highest LAMA3A message, while the uterus expresses the highest LAMA3B message 12. In situ RT-PCR of human embryonic tissues reveals positive staining for LAMA3 (not LAMA3A/B specific) message in developing tubules and developing comma-shaped bodies of the kidney, epithelial cells of the developing lung, in the basal layer of developing skin at gestational week 6.5, and in all layers of the epidermis from gestational week 8 onwards 19.…”

Laminin-311 (Laminin-6) Fiber Assembly by Type I-like Alveolar Cells

“…Thus, we have focused our studies on cultured primary alveolar cells. Although our previous work and that of others indicate that alveolar cells secrete a laminin-311 matrix, in this work we attempted to define the phenotype of the alveolar epithelial cell that deposits laminin-311 and to determine how alveolar cells organize laminin-311 into a fibrillar matrix (Miner et al 1997;Miosge et al 2002;Jones et al 2005).…”

“…For example, laminin a1 expression is restricted to early human lung morphogenesis, whereas laminin a3 and a5 expression are continuous from early lung development throughout adulthood (Virtanen et al 1996;Pierce et al 1998;Miosge et al 2002;Nguyen et al 2002b). Indeed, some authors have speculated that alveolar cells may express at least five laminin isoforms (Pierce et al 1998).…”