Junichi Shindo

Adhesion mediated by LFA-1 is required for efficient IL-12-induced NK and NKT cell cytotoxicity

Interleukin 12 (IL‐12)‐activated NK1.1+TCRα β+ (NKT2) and NK1.1+TCRα β– (NK) cells exhibit cytotoxic activity against a wide variety of tumor cells in the absence of prior sensitization. Here we demonstrate that the integrin adhesion receptor LFA‐1 (CD11a / CD18) regulates the cytotoxic activity of IL‐12‐activated NKT and NK cells against YAC‐1 and EL‐4 tumor cells. Differentiation in vivo and the expression of the cytolytic effector molecules perforin and Fas‐L were comparable in both IL‐12‐activated NKT and NK cells from LFA‐1– / – and LFA‐1+ / + mice. However, LFA‐1– / – IL‐12‐activated NKT and NK cells showed impaired conjugate formation with target cells. These results provide the first genetic evidence for a role for an adhesion receptor in killing by IL‐12‐activated NK cells.

Essential Role of the Adhesion Receptor LFA-1 for T Cell-Dependent Fulminant Hepatitis

Viral hepatitis affects more than 2 billion people worldwide. In particular, no effective treatment exists to abrogate death and liver damage in fulminant hepatitis. Activation of T cells is an initial and critical event in the pathogenesis of liver damage in autoimmune and viral hepatitis. The precise molecular mechanisms that induce T cell-mediated hepatocyte injury remain largely unclear. In mice, T cell-dependent hepatitis and acute liver damage can be modeled using ConA. In this study, we examined the role of the adhesion receptor LFA-1 in ConA-induced acute hepatic damage using LFA-1−/− (CD11a) mice. Massive liver cell apoptosis and metabolic liver damage were observed in LFA-1+/+ mice following ConA injection. By contrast, LFA-1−/− mice were completely resistant to ConA-induced hepatitis and none of the LFA-1−/− mice showed any hepatic damage. Whereas activated hepatic T cells remained in the liver in LFA-1+/+ mice, activated T cells were rapidly cleared from the livers of LFA-1−/− mice. Mechanistically, T cells from LFA-1−/− mice showed markedly reduced cytotoxicity toward liver cells as a result of impaired, activation-dependent adhesion. Importantly, adoptive transfer of hepatic T cells from LFA-1+/+ mice, but not from LFA-1−/− mice, sensitized LFA-1−/− mice to ConA-induced hepatitis. Thus, LFA-1 expression on T cells is necessary and sufficient for T cell-mediated liver damage in vivo. These results provide the first genetic evidence on an adhesion receptor, LFA-1, that has a crucial role in fulminant hepatitis. These genetic data identify LFA-1 as a potential key target for the treatment of T cell-mediated hepatitis and the prevention of liver damage.

Angiostatin gene therapy inhibits the growth of murine squamous cell carcinoma in vivo

Tumor growth is an angiogenesis-dependent process and therapeutic strategies aimed at inhibiting angiogenesis are theoretically attractive. Angiostatin has been shown to potently inhibit endothelial proliferation in vitro and tumor growth in vivo. We now show that a shift in the balance of tumor angiogenesis by gene transfer of a cDNA coding for mouse angiostatin into mouse squamous cell carcinoma NRS-1 and SCC-VII cells suppresses tumor growth in vivo. The inhibition of an angiostatin-transfected tumor was accompanied by a marked reduction in vascularity and the presence of many apoptotic tumor cells. However, transfected-angiostatin cDNA does not affect the expression of the vascular endothelial growth factor (VEGF) and VEGF-R2 in the vascular endothelium. The inhibition mechanisms of neovascularization may be mediated independent of VEGF:VEGF-R2 complex. Our data may provide a useful approach for human oral cancer therapy by gene therapy with angiostatin.

Myasthenia gravis associated with reduced masticatory function

This paper describes a case of myasthenia gravis in a 38-year-old woman who first consulted a dentist and then an oral surgeon because of chewing difficulty. Although myasthenia gravis commonly presents with diplopia, ptosis, or both as initial symptoms, chewing difficulty is rare. The patient was given steroid therapy and underwent thymectomy. Changes in bite force were monitored during treatment. The bite force was low when a high titer of anti-acetylcholine receptor antibody (11.0 nmol/l, normal <0.2) was found in the blood, but increased after the titer had decreased (1.5 nmol/l) in response to therapy.

Lidocaine action and conformational changes in cytoskeletal protein network in human red blood cells

The mechanism of action of lidocaine, which is commonly used clinically as a local anesthetic, was studied in human red blood cells. The influx of [14C]lidocaine through the cell membrane induced reversible transformation of human red blood cells from discocytes to stomatocytes. This change in shape depended on the lidocaine concentration and required both ATP and carbonic anhydrase. The lidocaine-induced shape change occurred as a result of spectrin aggregation, which altered the intracellular environment of the human red blood cells, mediated by carbonic anhydrase and activation of vacuolar type H(+)-ATPase (V-ATPase). Lidocaine controlled the influx of 22Na into the human red blood cells in a concentration-dependent manner. When incubated in media containing 6-chloro-9-[(4-diethylamino)-1-methyl-butyl]amino-2-methoxyacridine (mepacrine), an inhibitor of Na+ channels, human red blood cells changed shape from discocytes to stomatocytes and the intracellular pH decreased. This phenomenon was very similar to the shape change induced by lidocaine. These results suggest that the mode of action of lidocaine is related to a conformational change in the cytoskeletal protein network.

Growth of human squamous cell carcinoma xenografts in mice is inhibited by local angiostatin gene therapy

The possibility of inhibiting tumor growth by blocking the formation of new tumor vessels has recently received attention. Antiangiogenic tumor therapies have recently attracted intense interest because of their direct endothelial targeting and the absence of drug resistance. Local antiangiogenic gene therapy for cancer offers a potential way to achieve sustained therapeutic release of antiangiogenic substances. As a step toward this goal, we used liposomes complexed to angiostatin cDNA and targeted to human squamous cell carcinoma cell lines in vivo. Tumor cells expressing angiostatin after local gene transfer showed markedly reduced vascularity and contained many apoptotic tumor cells. These results demonstrate the potential utility of liposome-derived angiostatin for adjuvant therapy of oral cancer in humans.

Specific Identification of Telomerase Activity and p53 Gene Product in Metachronous Multiple Oral Squamous Cell Carcinoma and Adjacent Tissues

Abstract Objectives: To define the association of telomerase activity, histopathological characteristics, and p53 gene product expression in metachronous oral squamous cell carcinomas. Patients and Methods: Three regions of tissue specimens from the tumour, the region adjacent to the tumour, and intact oral mucosa from the surgical margin from 2 patients were simultaneously evaluated. Telomerase activity was determined by telomeric repeat amplification protocol assay. Hematoxylin and eosin stain and DO7 monoclonal antibody were used to stain formalin-fixed, paraffin-embedded sections to examine the histopathological features and p53 gene product expression. Results: Marked telomerase activity was expressed in tumour tissue and in adjacent dyskeratotic tissue. No telomerase activity was found in hyperkeratotic or dysplastic oral mucosa. Although p53 gene product was detected in the tumour and adjacent regions of both patients, it was found in the intact oral mucosa of only 1 patient. Conclusion: Estimation of tissue telomerase activity and p53 gene product expression is helpful in identifying malignancy in oral mucosal lesions.

The change of the immunological functions of cancer patients with the administration of interleukin-2

進行性口腔癌3症例に対しinterleukin-2 (IL-2) を投与して, 免疫学的活性を観察した。症例1: 77歳の男性, 頬粘膜再発性扁平上皮癌。2.59×107JRU (Japan Reference Unit) のIL-2を腫瘍周囲の組織内に24日間注射した。腫瘍の縮小が認められ, PRと診断したが, その後腫瘍は増殖し, 患者は4年後に死亡した。症例2: 66歳の女性, 頬粘膜の進行性扁平上皮癌。7.7×106JRUのIL-2を11日間静注した。腫瘍は増殖し, 放射線や化学療法を行ったが2か月後に死亡した。症例3: 63歳の男性, 上顎洞扁平上皮癌。上顎切除後, 翼突窩に腫瘍の残存が認められた。1.54×107JRUのIL-2静注が60Co照射と併用された。患者は1988年以来, 腫瘍を認めず生存中である。好酸球は症例1で増加し, リンパ球は症例1および3で増加した。数種類のリンパ球phenotypeは症例2で高値を示したが, 処置後は増加しなかった。症例1および3においてはCD57とCD16がIL-2投与後増加した。NKおよびLAK活性もまた投与後増大した。