Human placental laminin: Role in neuronal differentiation, cell adhesion and proliferation

Human placental extract has wound healing potential. Immuno-blots revealed presence of laminin in placentalextract (70 ± 0.257 lg/ml; n=3). It was purified using immuno-affinity chromatography. SDS-PAGE and SEHPLC indicated a188 kDa protein with some small peptides. Since placental laminin existed in its truncatedform, its roles in cellular migration, differentiation and wound healing were verified. Induction of cellularmigration and motility in rat fibroblasts were enhanced by placental laminin as observed from scratch woundassay. Promotion of neuronal differentiation of PC12 cells by placental laminin was observed by phase contrastmicroscopy. Confocal images showed presence of laminin on the cell surface and along the axonal processes.Significant interaction between integrin receptors and laminin responsible for cellular differentiation wasdemonstrated from co-localization experiments. Union between integrin receptor and its synthetic antagonistrevealed retarded pattern of neurite outgrowth in laminin treated cells. Animal model studies revealed fasterwound healing in the presence of placental laminin. Induction of re-epithelialization and angiogenesis inwound area by cellular proliferation and adhesion were observed. The cytokine levels showed an initial riseand gradual fall over the duration of wound healing on application of the fragmented laminin. Thus, roles ofplacental laminin in neuronal differentiation and wound healing were indicated.

Presence of bound substrate in lactate dehydrogenase from carp liver.

While attempting to purify UDP-galactose 4-epimerase from carp liver extract at pH 8.0, it was observed that the preparation even after dialysis could reduce NAD to NADH, interfering epimerase assay. The NAD reduction activity and the epimerase were co-eluted in a series of chromatographic steps. Mass spectrometric analysis of semi-purified fraction revealed that carp liver lactate dehydrogenase (LDH) contained bound lactate which was converted to pyruvate in the presence of NAD. The enzyme-bound lactate and the association with epimerase stabilized LDH from trypsin digestion and thermal inactivation at 45°C by factors of 2.7 and 4.2 respectively, as compared to substrate-free LDH. LDH and epimerase do not belong to any one pathway, but are the rate-limiting enzymes of two different pathways of carbohydrate metabolism. Typically, strongly associated enzymes work in combination, such as two enzymes of the same metabolic pathway. In that background, co-purification of LDH and epimerase as reloaded in this study was an unusual phenomenon.

Cell adhesion by aqueous extract of human placenta used as wound healer.

Aqueous extract of human placenta, used as wound healer, has shown significant cell adhesion property on mouse peritoneal macrophages and P388D1 cultured macrophage cell line. This property was offered primarily by fibronectin type III like peptide present in the extract and is comparable to fibronectin on a molar basis. The peptide induce adhesion of cell through cell surface receptors having K(d) = 2.8 +/- 0.9 x 10(-5) M suggesting weak binding. This is in support of integrins receptors that typically exhibit low affinities. Cell adhesion was partially inhibited by Arg-Gly-Asp (RGD) peptide, anti-beta1 integrin suggesting that integrin beta1 receptors have roles to play in the process.