Production of NGF by mouse hepatocellular carcinoma cells and expression of TrkA in tumor-associated arteries

W B Saunders, Kan, Kishibe; Yoshihisa, Yamada; Katsuhiro, Ogawa, GASTROENTEROLOGY, 122(7), 2002, 1978-1986 authorThe microarray analysis has revealed that NGF is specifically elevated in mouse hepatocellular carcinomas. The present study aimed to elucidate expression of NGF and its receptors (TrkA and p75NTR) during hepatic carcinogenesis, regeneration, development and primary hepatocyte culture in mice. Although NGF was negative or very weakly expressed in adult and developing livers, it was markedly elevated in neoplastic hepatocytic lesions from early stages of carcinogenesis. Appreciable levels were also detected in regenerating livers and hepatocyte cultures. The conditioned medium of hepatocellular carcinoma cells caused PC12 neurite outgrowth, but this was reduced with pretreatment of the conditioned medium with the anti-NGF antibody or NGF antisense expression in hepatocellular carcinoma cells. Although neither TrkA nor p75NTR was detectable in either hepatocellular carcinoma or normal hepatic cells, TrkA was demonstrated in the walls of tumor associated arteries that contain abundant nerve fibers. This study demonstrated that NGF is expressed by hepatocytes during carcinogeneis, regeneration and primary culture, but may have cells other than hepatocytes as the target. TrkA expression and abundance of nerve fibers in the walls of tumor-associated arteries suggest some possible role for NGF in angiogenesis

T-cell activation in tonsils of patients with pustulosis palmaris et plantaris

AuthorPustulosis palmaris et plantaris (PPP) is a well-known skin disease closely related to tonsillar focal infections, and tonsillectomy is very effective for treatment of this condition. However, etiology of PPP was unclear. In this study, we investigated the characteristics of tonsils from PPP patients by clinical, immunohistochemical, immunological, and molecular–biological approach, and considered the etiology of PPP. For 47 Japanese patients with PPP who have tonsillectomy in Asahikawa Medical College, the skin lesion of PPP was improved in 87% of PPP patients 12 months after tonsillectomy. In quantitative immunohistologic analysis by measurement of T-cell nodule areas on tonsillar sections from those patients, there was positive correlation between the enlargement and improvement of the skin lesion. These results suggested that the T-cell nodule expansion might be an important clue towards clarifying the pathogenesis of this tonsil related disease. In flow cytometric analysis, CD3+, CD4+ and CD4+CD25+ cells were significantly elevated in tonsillar lymphocyte from PPP patients. In reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting analyses of TGF-beta, IL-10, and Smad group, Smad7 mRNA and protein in tonsillar CD3+ cells from PPP patients were expressed at higher level than those from obstructive sleep apnea syndrome (OSAS) patients. These results suggested that helper T cells may be frequently activated and proliferating in tonsils of PPP patients, and the activation and proliferation of helper T cells may be due to enhanced inhibition by Smad7 in intracellular signal-transduction of TGF-beta

Expression of cutaneous lymphocyte-associated antigen (CLA) in tonsillar T-cells and its induction by in vitro stimulation with alpha-streptococci in patients with pustulosis palmaris et plantaris (PPP)

Academic Press, Nozawa, Hayabusa ; Kishibe, Kan ; Takahara, Miki ; Harabuchi, Yasuaki, Clinical Immunology, 116(1), 2005, 42-53. authorPustulosis palmaris et plantaris (PPP) is known to be a one of the tonsil-related diseases because tonsillectomy is quite effective in curing this condition. However etiological association between tonsils and PPP have not fully clarified yet. Cutaneous lymphocyte-associated antigen (CLA) is known to be a specific homing receptor that facilitates T-cell migration into skin. In this study, we investigated the expression of CLA on T-cells in tonsil, peripheral blood, and skin from patients with PPP. Two-color flow cytometric and two-color immunohistological analyses revealed that the numbers of CLA/CD3 double-positive cells in freshly isolated tonsillar mononuclear cells (TMC) and in tonsillar tissues were significantly higher in patients with PPP than in patients without PPP (P < 0.01, each). In vitro stimulus with α-streptococcal antigens enhanced CLA expression of tonsillar T-cells and TGF-β production of TMC in patients with PPP (P < 0.01, each), but did not in patients without PPP. In peripheral blood from PPP patients, the number of the CLA/CD3 double-positive cells significantly decreased at 6 months after tonsillectomy (P < 0.05). The CLA/CD3 double-positive cells and the postcapillary venule that expressed with a ligand of CLA, E-selectin, were found more frequently in the plantar skin from patients with PPP as compared to that from healthy volunteers (P < 0.01, each). These data suggest that a novel immune response to α-streptococci may enhance CLA expression on tonsillar T-cells through TGF-β production in patients with PPP, resulting in moving of CLA-positive tonsillar T-cells to skin and tissue damages. This may play a key role in pathogenesis of PPP

Increase of activated T-cells and up-regulation of Smad7 without elevation of TGF-beta expression in tonsils from patients with pustulosis palmaris et plantaris

authorPustulosis palmaris et plantaris (PPP) is known to be a skin disease related to tonsillitis, because the pustulosis often become exacerbated during acute tonsillitis and disappears after tonsillectomy. However, etiology of PPP remains unclear. In this study, we investigated the activation of tonsillar T-cell from PPP patients. Furthermore, we analyzed expressions of cytotoxic T-lymphocyte antigen-4 (CTLA4) that is a co-stimulatory molecule for inhibition of T-cell activation and of Smad7 that is a regulatory factor of TGF-beta intracellular signaling. For 47 Japanese patients with PPP who had tonsillectomy, the skin lesion was improved in 87% of PPP patient at 12 months after tonsillectomy. In quantitative immunohistologic analysis, T-cell nodules on tonsillar tissues from PPP patients were more expanded than those from the patients with obstructive sleep apnea syndrome (OSAS) (P = 0.015), and there was a positive correlation between the enlargement and clinical improvement (r = 0.422, P = 0.021). Flow cytometric analysis showed that the numbers of CD4+CD25+ and CD4+CD29+ cells in tonsils from PPP patients increased significantly compared to those from OSAS patients (P = 0.017, P = 0.016, respectively). Using reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting analyses with CD3+ tonsillar lymphocytes, we found that both expressions of Smad7 mRNA and protein were enhanced in PPP patients compared with OSAS patients (P = 0.03, P = 0.02, respectively), but expression of TGF-beta mRNA was not different between 2 groups. Although mRNA expression of CTLA4 was reduced in PPP patients compared with OSAS patients (P = 0.04), the CTLA4 surface protein expression was not different between 2 groups. These data suggest that helper T-cells are frequently activated in tonsils from PPP patients, and this activation may be related to unresponsiveness of TGF-beta1 by overexpression of Smad7. Such hyper-activation of T-cell may increase the risk of elicitation of self-reactive T-cell, being associated with pathogenesis of PPP

A study of costimulatory factors in tonsils of pustulosis palmaris et plantaris

Elsevier, Kan, Kishibe ; Miki, Takahara ; Hayabusa, Nozawa ; Yasuaki, Harabuchi, International Congress Series, 1257, 2003, 293-297. authorPustulosis palmaris et plantaris (PPP) is known to be closely related to tonsillar focal infections and tonsillectomy is quite effective. Hyper-immune response to alpha streptococci in patients with PPP was reported. On the other hand, costimulatory factors were studied on autoimmune disease. CD28 and cytotoxic T lymphocyte-associated 4 (CTLA4) participate in these factors. CD28 take part in T-cell activation, while CTLA4 take part in T-cell suppression. Therefore, we investigated CD28 and CTLA4 mRNA levels by reverse transcription-polymerase chain reaction (RT-PCR) in tonsillar tissues and CD3 positive lymphocytes from patients with PPP, recurrent tonsillitis (RT), and obstructive sleep apnea syndrome (OSAS). We also investigated the expression levels of CD28 and CTLA4 by flow cytometry analysis in tonsillar mononuclear cells stimulated with alpha streptococci in these patients. It was revealed that the CTLA4 mRNA in tonsillar CD3 positive lymphocytes from PPP patients was expressed at lower level than non-PPP patients. Then, the expression levels of CTLA4, comparison before and after stimulation, were lower level in PPP patients than non-PPP patients. These results suggest that the lower expression level of CTLA4 in PPP may cause hyper-immune response to alpha streptococci and abnormal regulation of T-cell activation

Neuropsin promotes oligodendrocyte death, demyelination and axonal degeneration after spinal cord injury

authorPrevious studies indicated that the expression of neuropsin, a serine protease, is induced in mature oligodendrocytes after injury to the central nervous system (CNS). The pathophysiology of spinal cord injury (SCI) involves primary and secondary mechanisms, the latter contributing further to permanent losses of function. To explore the role of neuropsin after SCI, histochemical and behavioral analyses were performed in wild-type (WT) and neuropsin-deficient (neuropsin^) mice using a crush injury model, a well-characterized and consistently reproducible model of SCI. In situ hybridization revealed that neuropsin mRNA expression was induced in the spinal cord white matter from WT mice after crush SCI, peaking at day 4. Neuropsin^ mice showed attenuated demyelination, oligodendrocyte death, and axonal damage after SCI. Although axonal degeneration in the corticospinal tract was obvious caudal to the lesion site in both strains of mice after SCI, the number of surviving nerve fibers caudal to the lesion was significantly larger in neuropsin^ mice than WT mice. Behavioral analysis revealed that the recovery at days 10-42 was significantly improved in neuropsin^ mice compared to WT mice in spite of the severe initial hindlimb impairments due to SCI in both strains. These observations suggest that neuropsin is involved in the secondary phase of the pathogenesis of SCI mediated by demyelination, oligodendrocyte death, and axonal degeneration