Accumulating data suggest that hepatic tolerance, initially demonstrated by spontaneous acceptance of liver allografts in many species, results from an immune regulatory activity occurring in the liver. However, the responsible cellular and molecular components have not been completely understood. We have recently described profound T cell inhibitory activity of hepatic stellate cells (HSCs) in vitro. In this study, we demonstrate in vivo evidence of immune modulatory activity of HSCs in mice using an islet transplantation model. Cotransplanted HSCs effectively protected islet allografts from rejection, forming a multilayered capsule, which reduced allograft immunocyte infiltrates by enhancement of apoptotic death. The immune modulation by HSCs appeared to be a local effect, and regulated by inducible expression of B7-H1, an inhibitory molecule of B7 family. This may reflect an intrinsic mechanism of immune inhibition mediated by liver-derived tissue cells. H epatic tolerance was initially recognized by spontaneous acceptance of liver allografts in a number of species. 1-4 Our laboratory focused on the mechanistic insights into liver transplant tolerance in mice and demonstrated that it is not attributable to a deletion of T cell clones specific to donor antigens, because lymphocytes isolated from tolerant mice respond well to donor antigen stimulation in vitro. 5 Whereas histological findings are notable for an abundance of CD4 and CD8 T cell infiltrates during the first week after transplantation, they are rapidly diminished via apoptotic death. [6][7][8] Similarly, in a nontransplant model, after injection of a specific antigenic peptide into TCR transgenic mice, deletion of specific CD8 ϩ T cells from the periphery was observed, as they accumulated in the liver and underwent subsequent apoptosis. 9 Combined, these results suggest a regulatory mechanism residing in the liver that may be responsible for its immune modulation. 10 However, the cellular components of the liver responsible for this are not completely understood.We have recently demonstrated that activated hepatic stellate cells (HSCs) effectively inhibit T cell responses in vitro, which is mediated by induction of T cell apoptosis. 11 Thus, subsequently studying whether HSCs can exert immune regulatory activities in vivo is logical. However, it is difficult to obtain direct evidence in the liver because approaches to specifically inhibit the effect of HSCs have not been defined. In this study, we used an islet allograft transplant model and demonstrated that cotransplantation with HSCs effectively protected islet allografts from rejection by forming a multi-layered immune barrier around the islets and reducing infiltrating T cells. Interestingly, HSCs isolated from B7-H1 knockout (KO) mice lost the protective effect on co-transplanted islet

Fibrosis is a common pathological sequela of tissue injury or inflammation, and is a major cause of organ failure. Subsets of fibroblasts contribute to tissue fibrosis in multiple ways, including generating contractile force to activate integrin-bound, latent TGFβ and secreting excess amounts of collagens and other extracellular matrix proteins (ECM) that make up pathologic scar. However, the precise fibroblast subsets that drive fibrosis have been poorly understood. In the absence of well-characterized markers, α-smooth muscle actin (αSMA) is often used to identify pathologic fibroblasts, and some authors have equated αSMA(+) cells with contractile myofibroblasts and proposed that these cells are the major source of ECM. Here, we investigated how well αSMA expression describes fibroblast subsets responsible for TGFβ activation and collagen production in three commonly used models of organ fibrosis that we previously reported could be inhibited by loss of αv integrins on all fibroblasts (using PDGFRβ-Cre). Interestingly, αSMA-directed deletion of αv integrins protected mice from CCl4-induced hepatic fibrosis, but not bleomycin-induced pulmonary or unilateral ureteral obstruction-induced renal fibrosis. Using Col-EGFP/αSMA-RFP dual reporter mice, we found that only a minority of collagen-producing cells coexpress αSMA in the fibrotic lung and kidney. Notably, Col-EGFP(+)αSMA-RFP(-) cells isolated from the fibrotic lung and kidney were equally capable of activating TGFβ as were Col-EGFP(+)αSMA-RFP(+) cells from the same organ, and this TGFβ activation was blocked by a TGFβ-blocking antibody and an inhibitor of nonmuscle myosin, respectively. Taken together, our results suggest that αSMA is an inconsistent marker of contractile and collagen-producing fibroblasts in murine experimental models of organ fibrosis.

Poly(l-lysine) (PLL) was end-grafted onto silicon oxide surfaces to study its conformational transition among α-helix, β-sheet, and random coil at the solid−water interface. Surface-grafted PLL films were prepared from surface-grafted poly(N ε -carbobenzyloxy-l-lysine) (PCBL) thin films, which were fabricated by the surface-initiated vapor-deposition polymerization (SI-VDP) of N-carboxyanhydride (NCA) of N ε -carbobenzyloxy-l-lysine. The reaction parameters during the SI-VDP process, including NCA concentration, substrate temperature, and reaction time, were optimized in order to synthesize homogeneous PCBL films with well-controlled thickness. Under the optimal conditions (substrate temperature at 105 °C, NCA evaporating temperature at 100 °C, and 8 mg of NCA at 0.1 Pa), α-helical PCBL films with thickness ranging from 4 to 120 nm can be synthesized by controlling the reaction time from 5 to 120 min. The conversion from PCBL to PLL was accomplished by removing N ε -carbobenzyloxy groups in the hydrogen bromide/benzene solution with the aid of ultrasonication for 40 min. The surface-grafted PLL films with an estimated DP of 750 exhibited versatile conformations among α-helix, β-sheet, and random coil depending on the external environments. The conformational transition from one state to another was successfully induced by external stimuli, such as pH (H+/OH-), surfactant (sodium dodecyl sulfate (SDS)), and anion (ClO4 -). In the process of the conformational transition, macroscopic properties such as thickness, refractive index, and wettability changed correspondingly. For example, a surface-grafted random-coiled PLL had a film thickness of 215 nm and a refractive index of 1.37 at pH 7, whereas it changed to a β-sheet conformation when binding with SDS, with the film thickness shrinking to 145 nm and the refractive index increasing to 1.45. Compared to the free PLL chains, the stability of the helical state increased when the PLL chains were densely immobilized on the surfaces. The transitional behaviors were characterized by Fourier transform infrared spectroscopy, circular dichroism spectroscopy, and ellipsometry.

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