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Biomedical subjects

H Ohguro

Publications and source records attributed to H Ohguro.

At least 19 recordsLinked to original sources
PubMed2001

Identification of human antitumor cytotoxic T lymphocytes epitopes of recoverin, a cancer-associated retinopathy antigen, possibly related with a better prognosis in a paraneoplastic syndrome.

Cancer-associated retinopathy (CAR) is a rare paraneoplastic syndrome, and the recoverin-specific autoantibody is suggested to contribute to the pathogenesis of retinopathy, including apoptosis of retinal cells. Because it is known that CAR(+) cancer patients have a preferable prognosis, we hypothesized that aberrantly expressed recoverin in cancer cells can become a target of cytotoxic T lymphocytes (CTL). Here we tested nine recoverin-derived HLA-A24-binding peptides for their capacity to elicit antitumor CTL. We observed recoverin-specific CTL responses in two HLA-A24(+) CAR(+) cancer patients. In addition, the CTL responses were obtained from three of ten CAR(-) cancer patients and two of six healthy individuals. The CTL precursor frequency of CAR(+) cancer patients and that of CAR(-) cancer patients was higher than that of healthy individuals. Of nine recoverin peptides, R49 (QFQSIYAKF), R49.2 (QFQSIYAKFF), and R64 (AYAQHVFRSF) were discovered to induce the peptide-specific CTL. Taken together, our present data suggest that peripheral activation of recoverin-specific antitumor CTL is likely to contribute to the preferable prognosis of CAR(+) cancer patients. Moreover, in cases other than CAR(+) cancer patients, recoverin may offer the opportunity to design epitope-based immunotherapeutic approaches for treating HLA-A24(+) cancer patients with a recoverin-expressing tumor.

Antigens, Neoplasm↗
PubMed2001

Mechanisms of photoreceptor cell death in cancer-associated retinopathy.

PURPOSE: In a previous study, both recoverin and heat shock cognate protein (hsc) 70 were recognized as autoantigens by sera from patients with cancer-associated retinopathy (CAR), and retinal dysfunction similar to CAR was inducible by intravitreous injection of anti-recoverin and anti-hsc 70 antibodies to Lewis rat. The purpose of the present study was to elucidate the effects of these antibodies on retinal photoreceptor cell functions, the contribution of caspase during the photoreceptor degeneration, and the roles of aberrant expression of recoverin in tumor cells. METHODS: As photoreceptor functions, rhodopsin phosphorylation using freshly prepared rod outer segments (ROS) and electroretinogram (ERG) were studied. Expression of recoverin in several kinds of tumors was examined by reverse transcription-polymerase chain reaction and Western blot analysis. The effects of recoverin on calcium-dependent protein phosphorylation were studied using the A549 lung adenocarcinoma cell line, which does not express recoverin. RESULTS: Rhodopsin phosphorylation in bovine ROS was significantly promoted by the addition of anti-recoverin antibody. Similar effects on rhodopsin phosphorylation and ERG impairment were observed in rat eyes treated with anti-recoverin antibody. Co-injection of caspase inhibitors with anti-recoverin antibody inhibited ERG impairment and significantly suppressed the antibody-induced enhancement of rhodopsin phosphorylation. Aberrant expression of recoverin was found in 15 of 30 tumor tissues from patients with cancer without CAR. Profiles of calcium-dependent protein phosphorylation of cell lysate from A549 cells were modulated by the presence of purified recoverin. CONCLUSIONS: These observations suggest that anti-recoverin antibody is incorporated into rod photoreceptor cells and modulates rhodopsin phosphorylation, which in turn produces activation of caspase-dependent apoptotic pathways. Regarding antibody generation in CAR, a high incidence of aberrant expression of recoverin in cancer tissues is important, as suggested previously.

Animals↗
PubMed2001

[Autoimmune mechanisms in molecular pathology of glaucomatous optic neuropathy].

BACKGROUND: Glaucomatous optic neuropathy is characterized by loss of retinal ganglion cells and their axons, excavated appearance of the optic nerve head, and progressive loss of visual field sensitivity. In terms of pathology of the glaucomatous optic neuropathy, observations suggest retinal ganglion cell death by apoptosis. Deprivation of neurotrophic factors, ischemia, chronic elevation of glutamate, and disorganized nitric oxide metabolism have been suspected to be mechanisms triggering apoptosis. Another recent suggestion is that autoantibodies directed toward retinal antigens may be involved in facilitating apoptotic cell death in some glaucoma patients. METHOD: We summarized previous reports dealing with the relationship between glaucoma etiology and autoimmune responses, and investigated the contribution of autoimmune factors in glaucomatous optic neuropathy using electrophysiological and biochemical methods. RESULTS: We found serum anti-neuron specific enolase(NSE) antibody in approximately 20% of our glaucoma patients. In primary open angle glaucoma, maximum and mean intraocular pressures in antibody positive patients were significantly or relatively lower than pressures in antibody negative patients. In addition, we also found that apoptotic cell death of retinal ganglion cells was induced in rat eyes by intravitreal administration of purified anti-NSE antibody. CONCLUSION: As suggested by previous reports, autoimmune response is one of the mechanisms related to glaucoma etiology and NSE may have an important function as an autoantigen.

Animals↗
PubMed2001

Retinal dysfunction in cancer-associated retinopathy is improved by Ca(2+) antagonist administration and dark adaptation.

PURPOSE: It was recently found that recoverin acts as an autoantigen recognized by sera of patients with cancer-associated retinopathy (CAR), and that CAR-like retinal dysfunction is produced by intravitreous administration of anti-recoverin antibody in Lewis rat eyes. To examine the pathologic molecular mechanism of CAR, and to elucidate an effective therapy for CAR, the function and morphology of CAR were compared with those of phototoxic retinal damage, another form of photoreceptor dysfunction, and the effect of nilvadipine, a Ca(2+) antagonist, on the retinal degenerations was studied, using these models. METHODS: Under different illumination conditions and/or medication with nilvadipine, the functional and morphologic properties of the retinas were evaluated after intravitreous injection of anti-recoverin antibody into Lewis rat eyes (six rats, 12 eyes in each experimental condition), using electroretinogram (ERG), rhodopsin phosphorylation, and light microscopy. RESULTS: Anti-recoverin antibody administered into the vitreous of Lewis rat eyes induced a significant decrease and increase of ERG responses and rhodopsin phosphorylation levels, respectively, under cyclic or continuous light. Similar changes were observed in eyes of rats bred under continuous illumination that did not receive anti-recoverin antibodies. However, anti-recoverin antibody-induced retinal dysfunctions were not observed in rat eyes under dark conditions. Administration of nilvadipine, a Ca(2+) antagonist, to the anti-recoverin antibody-treated rats and rats with phototoxic retinal dysfunction caused significant improvement of the deterioration of ERG and normalization of rhodopsin phosphorylation. CONCLUSIONS: The present data indicate that anti-recoverin antibody-induced retinal dysfunction was functionally similar to phototoxic retinal dysfunction and was markedly suppressed under dark conditions or by systemic administration of a Ca(2+) antagonist.

Animals↗
PubMed2000

Aberrant expression of photoreceptor-specific calcium-binding protein (recoverin) in cancer cell lines.

Cancer-associated retinopathy (CAR) is an ocular manifestation of a paraneoplastic syndrome whereby immunological reactions to retinal antigens aberrantly expressed in tumor cells lead to the degeneration of retinal photoreceptor cells. In our previous study (H. Ohguro et al., Invest. Ophthalmol. Vis. Sci., 40: 82-89, 1999), recoverin, a retina-specific calcium-binding protein, and heat shock cognate protein 70 (hsc 70) were identified as autoantigens recognized by sera from patients with CAR. Therefore, we suggested that autoimmune reactions against both recoverin and hsc 70 might be involved in the pathogenesis of CAR. To elucidate the initial step of the molecular pathology of CAR, we examined the expression of recoverin and hsc 70 by reverse transcription-PCR and Western blot using cell lines of several kinds of cancers, including lung small cell carcinoma, lung adenocarcinoma, gastric cancer, pancreatic cancer, breast cancer, uterine cervical cancer, endometrial cancer, and leukemia. Recoverin was expressed in 21 of the 31 cancer cell lines. The expression levels of hsc 70 were significantly higher in cancer cell lines than in noncancerous cell lines. However, no difference in the expression levels of hsc 70 was observed between recoverin-positive and -negative cell lines. Immunofluorescence labeling by the affinity-purified recoverin antibody revealed the immunoreactivity to recoverin as a granular pattern within the cancer cells. Lung adenocarcinoma A549 cells, which did not express recoverin, exhibited a significant reduction in cell proliferation upon transfection with human recoverin cDNA. Taken together, our present data suggest that the retina-specific calcium-binding protein recoverin is expressed in more than 50% of a variety of cancer cells and may play a significant role in the cell proliferation of these tumor cells.

Antigens, Neoplasm↗
PubMed2000

Purification and characterization of bovine cone arrestin (cArr).

To elucidate the quenching mechanism of phototransduction in vertebrate cone photoreceptors, a cDNA clone encoding cone specific arrestin (cArr) was isolated from a bovine retinal cDNA library using a human cArr cDNA probe. Affinity-purified anti-peptide antibody specific to cArr was prepared. Immunohistochemical staining displayed specific labeling of cArr in cone photoreceptors and immunoblotting identified a 46 kDa protein band. We purified cArr from bovine retinas by sequential column chromatography using DEAE-cellulose, gel filtration and mono Q columns. Binding studies revealed no binding of cArr to rhodopsin regardless of whether it was bleached and/or phosphorylated. cArr also failed to bind to heparin-Sepharose under conditions which rod arrestin (rArr) bound to the column. The present data suggest that cArr may play a role in the quenching of phototransduction in cone photoreceptors and that its activity therein is different to that of rArr.

Amino Acid Sequence↗
PubMed2000

Retinal ganglion cells recognized by serum autoantibody against gamma-enolase found in glaucoma patients.

PURPOSE: To study pathologic roles of the presence of serum autoantibodies against retinal ganglion cells in patients with glaucoma. METHODS: Serum autoantibody reactions were detected by Western blot analysis using retinal soluble fractions in 79 patients with glaucoma (normal-tension glaucoma [NTG], 23 cases; primary open-angle glaucoma [POAG], 56 cases) and 60 age-matched healthy subjects. Clinical characteristics including visual acuity, visual field, intraocular pressure (IOP), and optic disc features were compared between the serum autoantibody-positive and -negative patients. The retinal autoantigen recognized by patients' sera was identified by a combination of in-gel digestion and Edman sequencing. RESULTS: Western blot analysis revealed that serum autoantibody against retinal 50-kDa antigen was recognized in 20 out of 79 glaucoma patients (25.3%; 14 POAG and 6 NTG patients) and 60 age-matched control subjects (11.7%), respectively. Immunocytochemistry revealed that labeling of the ganglion cell layer (GCL) by IgG from glaucoma patients (POAG: 13/56, 23.2%; NTG: 6/23, 26%) existed at a significantly higher rate than that by IgG from control subjects (2/60, 3.3%; P < 0.05). In POAG, maximum IOP in the serum antibody positive-patients was significantly lower than that in the antibody-negative patients (P < 0.05). However, no statistical differences were observed in visual field loss, disc cupping, and other clinical factors between the antibody-positive and -negative groups in POAG and NTG. In-gel digestion of the 50-kDa band in two-dimensional polyacrylamide gels and Edman sequence analysis of the high-performance liquid chromatography-purified peptides identified the 50-kDa protein as gamma-enolase. Injection of the 50-kDa IgG from glaucoma patients or anti-gamma-enolase serum into the vitreous cavity of Lewis rats caused reduction of the b-wave of the electroretinogram and TdT-dUTP terminal nick-end labeling (TUNEL)-positive staining within the GCL. CONCLUSIONS: In the current study, serum autoantibody against 50-kDa protein identified as gamma-enolase in 25% of glaucoma patients.

Aged↗
PubMed1999

Detection of serum antibody against arrestin from patients with acute disseminated encephalomyelitis.

In our previous study, we found the presence of serum autoantibody against arrestin in patients with multiple sclerosis (MS), while such serum autoantibody was not detected from patients with other neurological diseases and control subjects. We suggested that serum arrestin antibody titers may be useful for the diagnosis and evaluation of the disease's course. In the present study we examined sera from 7 patients, who were initially diagnosed as having acute disseminated encephalomyelitis (ADEM), for the presence of serum antibody against arrestin, in order to study the specificity of the serum antibody among demyelinated diseases. High titers were detected from 2 patients out of 7. One of the patients, a 4 year-old girl, presented with an additional neurological attack during the 6 months after the initial attack, resulting in change of diagnosis to MS. During her disease course the serum titers against arrestin fluctuated in correspondence with the disease's activity. These observations suggest that the presence of serum autoantibody against arrestin may be specific to MS and be helpful for differential diagnosis of ADEM and MS.

Adolescent↗
PubMed1999

Low expression of alphaA-crystallins and rhodopsin kinase of photoreceptors in retinal dystrophy rat.

PURPOSE: The Royal College of Surgeons (RCS) rat has been extensively characterized as a model for inherited retinal dystrophy such as retinitis pigmentosa. In the present study, compositions of retinal proteins were compared between RCS (rdy-/-) and control (rdy+/+) rats during progression of the disease to understand the molecular pathologic course of the retinal degeneration. METHODS: Protein mapping was performed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) or two-dimensional (2D)-PAGE using whole retinas or rod outer segments (ROS) obtained by a sucrose-density gradient centrifugation method from RCS or control rats at the age of 3 to 8 weeks. RESULTS: 2D-PAGE showed that retinal proteins of RCS rats were generally less abundant than those of the control animals and that the difference became more evident with aging. However, no significant difference was observed in the protein-mapping patterns in 2D-PAGE between RCS and control rats in any ages tested. Analysis by SDS-PAGE of ROS proteins and by western blot using antibodies against opsin, rhodopsin kinase (RK), recoverin, or arrestin demonstrated that a 20-kDa protein and RK were selectively less abundant in RCS than in control rats. Edman sequence analysis of the proteolytic peptides obtained by in-gel digestion of the corresponding protein band using endoproteinase Lys C identified the 20-kDa protein as alphaA-crystallin. Reverse transcription-polymerase chain reaction confirmed selective low levels of mRNA expressions of alphaA-crystallins and RK in RCS rats. CONCLUSIONS: This study demonstrates that decreased expression of alphaA-crystallins and RK in RCS rats, may have significant roles in the development of retinal dystrophy.

Amino Acid Sequence↗
PubMed1999

Cancer-associated retinopathy induced by both anti-recoverin and anti-hsc70 antibodies in vivo.

PURPOSE: In a previous study, both recoverin and heat shock cognate protein 70 (hsc 70) were found as autoantigens recognized by sera from four patients with cancer-associated retinopathy (CAR). This observation suggested that autoimmune reactions against recoverin and hsc 70 might be involved together in the pathogenesis of CAR. The purpose of the present study is to investigate the effects of these autoantibodies on retinas in vivo. METHODS: Functional and morphologic properties of the retinas were evaluated after anti-recoverin and/or anti-hsc 70 antibodies were intravitreously injected into Lewis rats' eyes. RESULTS: Responses in electroretinogram (ERG) of eyes penetrated with anti-hsc 70 antibody were comparable with the control, but those with anti-recoverin antibody were remarkably reduced during the 3-week period after the injection. Such anti-recoverin antibody-induced reduction was significantly enhanced by copenetration with anti-hsc 70 antibody. Immunofluorescence microscopy demonstrated that after intravitreal injection, anti-recoverin antibody penetrated toward the outer nuclear layer (ONL) and outer segments within 12 to 24 hours, and the presence of the antibody in the retina diminished during the next few days. Histopathology revealed significant thinning of the ONL and inner nuclear layer (INL) in the affected retina in comparison with the control. Throughout the ONL and INL, apoptotic cells were recognized by TdT-dUTP terminal nick-end labeling. The antibody-induced retinal dysfunction was effectively treated by administrations of either corticosteroid or cyclosporin A. CONCLUSIONS: These observations suggest that anti-recoverin- and anti-hsc 70 antibody-induced retinal dysfunction in Lewis rat is a good model to study the pathophysiology of CAR.

Animals↗
PubMed1999

Recoverin and Hsc 70 are found as autoantigens in patients with cancer-associated retinopathy.

PURPOSE: To characterize retinal autoantigens in patients with cancer-associated retinopathy (CAR). METHODS: The sera of 4 patients with CAR were examined by western blot analysis, and the proteins specifically probed were partially purified from bovine retinas and identified by Edman sequence analysis of the proteolytic peptides. RESULTS: Western blot analysis demonstrated that soluble 23-kDa and 65-kDa proteins were probed by the serum of all the patients. The 23-kDa protein assumed an identical position to that of recoverin when the latter, previously identified as an autoantigen of CAR, was probed by its antibody. This strongly suggested that the 23-kDa antigen and recoverin are identical proteins. After partial purification of the 65-kDa protein from bovine retinas, the corresponding band in two-dimensional gel electrophoresis was cut out and subjected to in-gel digestion by endoproteinase Lys C. Edman sequencing of the proteolytic peptides purified on a high-performance liquid chromatography reverse-phase column identified the 65-kDa protein as the heat shock cognate protein 70 (hsc 70), a member of the heat shock protein (hsp) family involved in protein metabolism as chaperons under both stress and nonstress conditions. To estimate the relationship between the autoimmune responses against recoverin and hsc 70, the maximum serum dilutions required to identify recoverin and hsc 70 on western blot analysis were determined and found to be different among the patients. CONCLUSIONS: These observations suggest that humoral autoimmune reactions against recoverin and hsc 70 might be involved in the pathogenesis of CAR.

Adenocarcinoma↗
PubMed1998

Characterization of multi-input electroretinogram using normal control subjects.

BACKGROUND: The multi-input electroretinogram (ERG) has recently been developed to analyze focal retinal responses in the central region and is expected to become a powerful tool for management of retinal diseases. The relationship between the obtained ERG response and retinal neuronal functions is not yet fully understood. METHOD: In the present study, in order to study the spatial retinal functions; multi-input ERGs were recorded from 14 healthy subjects under several illumination conditions modulating photoreceptor functions. RESULTS: The first-order kernel amplitudes were changed in correspondence to the cone photoreceptor adaptation states under different background illuminations. A strong flash exposure significantly lowered the amplitude (approximately 70%) within the central 15 deg but lowered it only slightly (10-20%) in the peripheral areas outside the central 15 deg. Similar effects were observed when the stimulus luminance was lowered by neutral density filters. CONCLUSION: Our data suggest that a newly developed multi-input measurement of ERGs represents signals within the firing of postreceptor neurons from cone photoreceptors at specific areas. Thus, this analysis is a useful tool for mapping the spatial network of neurons in the central region.

Adult↗
PubMed1998

A case of a combination of Oguchi's disease and congenital retinoschisis.

We report here a rare case of a combination of Oguchi's disease and congenital retinoschisis. A 36-year-old male patient presented with a decrease in vision in both eyes and night blindness. Indirect ophthalmoscopy revealed bilateral macular stellate striations and golden-gray discoloration of the retina outside the vascular arcades. This discoloration turned to a normal retina after complete adaptation to darkness (Mizuo-Nakamura phenomenon).

Adult↗
PubMed1998

Identification of a single phosphorylation site within octopus rhodopsin.

Light-dependent phosphorylation of rhodopsin (Rho) is a first step in the desensitization of the signaling state of the receptor during vertebrate and invertebrate visual transduction. We found that only 358Ser of the photoactivated octopus Rho (oRho*) was phosphorylated by octopus rhodopsin kinase (oRK). Tryptic truncation of the C-terminal PPQGY repeats of oRho that follow the phosphorylation region did not influence spectral or G-protein activation properties of oRho but abolished phosphorylation. Despite significant structural differences between oRK and mammalian RK, these results provide further evidence of the importance of singly phosphorylated species of Rho* in the generation of arrestin binding sites.

Amino Acid Sequence↗