Basic fibroblast growth factor stimulates the proliferation of human dermal fibroblasts via the ERK1/2 and JNK pathways.

BACKGROUND: Basic fibroblast growth factor (bFGF, FGF-2) has been described as a multipotent cytokine that regulates cell growth as well as differentiation, matrix composition, chemotaxis, cell adhesion and migration in numerous cell types. It is known that bFGF stimulates proliferation of cultured fibroblasts. However, the detailed mechanism of fibroblast proliferation induced by bFGF in vitro still remains to be elucidated. Objectives We investigated the precise effects of bFGF on fibroblast proliferation and the signalling pathways responsible for bFGF-induced proliferation in cultured human dermal fibroblasts (HDFs). METHODS: HDFs were cultured with bFGF in the presence or absence of specific inhibitors against MAPK signalling pathways including ERK, JNK and p38. The number of cells was counted and immunoblotting findings were examined for the activation of ERK1/2 and JNK. Furthermore, the inhibitory effects of ERK1, ERK2 and JNK1 were proven by the transfection of siRNA. RESULTS: bFGF increased the number of HDFs in a dose- and time-dependent manner. The bFGF-induced proliferation was suppressed by the MEK inhibitors PD98059 and U0126, and the JNK inhibitor SP600125. bFGF increased the phosphorylation levels of ERK1/2 and JNK1. Treatment with ERK1, ERK2 or JNK1 siRNA significantly inhibited bFGF-induced proliferation. CONCLUSIONS: This study indicates that ERK1/2 and JNK pathways play an important role in the bFGF-mediated effect in HDFs. This study also suggests that controlling ERK1/2 and/or JNK signalling may therefore be a new therapeutic approach for the treatment of chronic and untreatable skin ulcers.

Dissociation of ligand-induced internalization of CXCR-4 from its co-receptor activity for HIV-1 Env-mediated membrane fusion.

The C-terminal cytoplasmic tail of chemokine receptors is important for their internalization upon ligand binding. We generated several deletion mutants of the C-terminal cytoplasmic tail of CXCR-4, a co-receptor for T cell line tropic strains of human immunodeficiency virus type 1 (HIV-1), to know whether or not co-receptor internalization is associated with HIV-1 entry. Our data showed that the removal of C-terminal 15 amino acid residues of the cytoplasmic tail from CXCR-4 completely abolished its internalization, but did not affect the co-receptor activity at all. Co-receptor activity was fully retained even when all 45 amino acid residues in the C-terminal cytoplasmic tail had been deleted. These data indicated that no cytoplasmic tail nor internalization of CXCR-4 is required for its co-receptor activity for HIV-1 entry.

Large quantity production with extreme convenience of human SDF-1alpha and SDF-1beta by a Sendai virus vector.

We describe a robust expression of human stromal cell-derived factor-1alpha (SDF-1alpha) and SDF-1beta, the members of CXC-chemokine family, with a novel vector system based upon Sendai virus, a non-segmented negative strand RNA virus. Recombinant SDF-1alpha and SDF-1beta were detected as a major protein species in culture supernatants, reached as high as 10 microg/ ml. This remarkable enrichment of the products allowed us to use even the crude supernatants as the source for biological and antiviral assays without further concentration nor purification and will thus greatly facilitate to screen their genetically engineered derivatives.

Infectivities of human and other primate lentiviruses are activated by desialylation of the virion surface.

The envelope protein, gp120, of human immunodeficiency virus type 1 (HIV-1) is heavily glycosylated and sialylated. The heavy sialylation greatly affects the physical properties of the protein, as it resolves into a wide acidic pH range despite the basic pI value predicted for its polypeptide backbone (B. S. Stein and E. G. Engleman, J. Biol. Chem. 265:2640-2649, 1990). However, the functional significance of the heavy sialylation remains elusive. Here, we show that desialylation of HIV-1 with neuraminidase greatly augments the initial virus-cell interaction, leading to remarkably enhanced viral replication and cytopathogenicity. This enhancement appeared to be a direct result of the removal of negatively charged sialic acids but not of the exposure of galactose residues or complement activation. Complementing these results, studies with inhibitors of mannosidase I and mannosidase II showed that the processing of HIV-1 oligosaccharides into the complex type to acquire the terminal sialic acid residues impeded the full replication capacity of the virus and that its prevention also enhanced virus replication and cytopathogenicity. Enhancement of infection by desialylation was found widely, with HIV-1 laboratory strains of different cell tropisms and primary isolates as well as HIV-2 and simian immunodeficiency virus. Thus, the sialylation catalyzed by host cell pathways appeared to reduce the infectivity of human and nonhuman primate lentiviruses. Our results further suggested that desialylation would help increase the titers of HIV-based vectors.

Review of studies on the immunological capacity in the bursectomized chick.

This paper summarizes data on the immunological capacity in the bursectomized chick. A series of experiments described by Glick and Sadler represented the functional importance of the bursa of Fabricius for the humoral immunity in chicken. Later studies of immune responses in bursaless chickens reported by Lerner et al. contributed to our knowledge of bursa-independent humoral immunity and demonstrated an extra-bursal site for B-cell differentiation. Bursectomy at an early stage of chicken development changes the immune responses after hatching. Here I present my current understanding of embryonic B-cell populations (bursa-dependent and independent) following in ovo bursectomy which may influence B-cell differentiation with reference to our experiments on J chain production.