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Biomedical subjects

Takayuki Harada

Publications and source records attributed to Takayuki Harada.

46 records · Page 3Linked to original sources
PubMed2002

Microglia-Müller glia cell interactions control neurotrophic factor production during light-induced retinal degeneration.

Activation of microglia commonly occurs in response to a wide variety of pathological stimuli including trauma, axotomy, ischemia, and degeneration in the CNS. In the retina, prolonged or high-intensity exposure to visible light leads to photoreceptor cell apoptosis. In such a light-reared retina, we found that activated microglia invade the degenerating photoreceptor layer and alter expression of neurotrophic factors such as nerve growth factor (NGF), ciliary neurotrophic factor (CNTF), and glial cell line-derived neurotrophic factor (GDNF). Because these neurotrophic factors modulate secondary trophic factor expression in Müller glial cells, microglia-Müller glia cell interaction may contribute to protection of photoreceptors or increase photoreceptor apoptosis. In the present study, we demonstrate the possibility that such functional glia-glia interactions constitute the key mechanism by which microglia-derived NGF, brain-derived neurotrophic factor (BDNF), and CNTF indirectly influence photoreceptor survival, although the receptors for these neurotrophic factors are absent from photoreceptors, by modulating basic fibroblast growth factor (bFGF) and GDNF production and release from Müller glia. These observations suggest that microglia regulate the microglia-Müller glia-photoreceptor network that serves as a trophic factor-controlling system during retinal degeneration.

Animals↗
PubMed2002

Role of nucleotide sequences in the V3 region in efficient replication of CCR5-utilizing human immunodeficiency virus type 1 in macrophages.

Macrophages express both CXCR4 and CCR5 coreceptors, but restrict X4 HIV-1 replication unless the Env-V3 region, a major determinant of cell tropism, is exchanged with that of R5 HIV-1. As the V3 exchange concomitantly alters the nucleotide sequences, we introduced silent mutations in the V3 or C2 region of macrophage-tropic R5 JRFL without changing the amino acids. Immunoblot analysis confirmed that viral proteins including Env-gp120 were similarly incorporated in wild-type (wt) and mutant virions. The silent mutants infected CCR5-positive MAGIC5 cells but not CCR5-negative MAGI cells, as productively as wt viruses, indicating that the silent mutations did not alter coreceptor utilization. In contrast, two of three silent V3-mutant viruses failed to replicate efficiently in primary macrophages, whereas other V3- or C2-mutants and wt JRFL infected macrophages productively. Furthermore, synthesis of the full-length viral DNA of the aberrant V3-mutant was largely reduced in macrophages. These results suggest that V3 nucleotide sequences may be one of the postentry factors restricting HIV-1 replication in macrophages.

Base Sequence↗
PubMed2002

MD41, a novel T helper 0 clone, mediates mast-cell dependent delayed-type hypersensitivity in mice.

In a previous study on mouse, we have shown that delayed-type hypersensitivity (DTH) could be classified into two types according to MC requirement. The first type of DTH could be elicited by sensitization with methylated human serum albumin (MHSA) in complete Freund's adjuvant (CFA) in both wild type and mast-cell deficient (W/Wv) mice. The second type could be elicited by MHSA in incomplete Freund's adjuvant (IFA) sensitization in wild type but not W/Wv mice. While the former was related to classic tuberculin (tbc)-type DTH, the latter appeared to be a novel mast-cell dependent DTH (MD-DTH). In order to investigate the mechanism of MD-DTH, in this study, we generated an effector T-cell clone (MD41) from lymph node cells of MHSA in IFA-sensitized mice and analysed its pattern of cytokine production. Our results from cytokine assays show that following antigen stimulation, MD41 cells produce significant amounts of the T helper 1 (Th1) cytokine interferon-gamma (IFN-gamma) as well as the Th2 cytokines interleukin (IL)-4 and IL-10. In addition, double staining for IL-4 and IFN-gamma revealed that MD41 cells produce both Th1- and Th2-type cytokines simultaneously, which suggest that MD41 represents a Th0 clone rather than a mixture of Th1 and Th2 clones. Adoptive transfer of MD41 cells into wild-type mice resulted in the development of DTH skin reactions similar to those produced by active sensitization, with very similar histological findings. However, DTH skin reactions could not be induced in W/Wv mice unless first reconstituted with normal bone marrow MC (BM-MC). Therefore, our study suggests that in conjunction with tissue MC, MD41, a less-polarized MD-DTH-derived Th0 clone, is capable of developing murine DTH to the same extent as strongly polarized Th1 cells and mediates MD-DTH rather than tbc-type DTH.

Adoptive Transfer↗
PubMed2002

Neurotrophic factor receptors in epiretinal membranes after human diabetic retinopathy.

OBJECTIVE: Formation of epiretinal membranes (ERMs) in the posterior fundus results in progressive deterioration of vision. ERMs have been associated with numerous clinical conditions, including proliferative diabetic retinopathy (PDR), but its pathogenic mechanisms are still unknown. This study was conducted to determine whether neurotrophic factor receptors (tyrosine kinase receptors trkA, trkB, and trkC; low-affinity neurotrophin [NT] receptor p75 [p75(NTR)]; glial cell line-derived neurotrophic factor receptor-alpha1 [GFR alpha 1] and GFR alpha 2; and Ret) are involved in the formation of ERMs after PDR. RESEARCH DESIGN AND METHODS: ERM samples were obtained by vitrectomy from 19 subjects with PDR aged 57 +/- 8 years with 17 +/- 8 years of diabetes and 15 subjects with idiopathic ERM. They were processed for RT-PCR analysis. In addition, 11 ERM samples from PDR patients aged 47 +/- 18 years with 13 +/- 4 years of diabetes were processed for immunohistochemical analysis. RESULTS: Expressions of trkA, trkB, trkC, p75(NTR), and Ret mRNAs were similar in both groups. In contrast, GFR alpha 2 expression levels were significantly higher (17 of 19 vs. 2 of 15 subjects in idiopathic ERM, P < 0.0001) in PDR subjects. Accordingly, immunohistochemical analysis revealed expression of GFR alpha 2 protein in all of the 11 ERMs derived from PDR patients, and that region was double-labeled with glial cell-specific markers. On the other hand, GFR alpha 1 expression was lower (8 of 19 vs. 12 of 15 subjects with idiopathic ERM, P = 0.0258) in PDR subjects. CONCLUSIONS: These results suggest a possibility that glial cell line-derived neurotrophic factor receptor (GDNF) subtypes are differently involved in the formation of ERMs.

Adult↗
PubMed2002

Involvement of p27(KIP1) degradation by Skp2 in the regulation of proliferation in response to wounding of corneal epithelium.

PURPOSE: To examine the expression of the p27(KIP1)in the normal and epithelial-scraped cornea and whether degradation of p27(KIP1)by Skp2 is involved in the regulation of cell proliferation in response to wounding of the corneal epithelium. METHODS: C57Bl6, p27(KIP1-/-), Skp2(-/-), and Skp2(-/-)/p27(KIP1-/-) double-knockout mice were examined. Normal and epithelial-scraped corneas were analyzed by immunocytochemistry using anti-p27(KIP1) antibody. Cells in the S phase of DNA synthesis were analyzed by immunocytochemistry using anti-bromodeoxyuridine (BrdU) antibody. RESULTS: The p27(KIP1) was expressed in basal cells of the central and peripheral region of the cornea and limbus. This expression was not detected 24 hours after the epithelial scraping, when there were many cells in the S phase of DNA synthesis in the corneal epithelium. There were no obvious differences in the thickness and anti-BrdU staining in the corneal epithelium of p27(KIP1-/-) mice from that of control animals. Twenty-four hours after epithelial scraping in the Skp2(-/-) mice, the corneal epithelium was thinner than in wild-type mice and had many p27(KIP1)-positive cells and few BrdU-positive cells. In contrast, 24 hours after epithelial scraping in the Skp2(-/-)/p27(KIP1)(-/-) double-knockout mice, the corneal epithelium was as thick as in wild-type mice and had many BrdU-positive cells. CONCLUSIONS: These results suggest that degradation of p27(KIP1) by Skp2 is involved in the regulation of cell proliferation in response to wounding of the corneal epithelium.

Animals↗
PubMed2002

Amino-terminal phosphorylation of c-Jun regulates apoptosis in the retinal ganglion cells by optic nerve transection.

PURPOSE: To examine the involvement of c-Jun and c-Jun N-terminal phosphorylation (JNP) in apoptosis of retinal ganglion cells (RGCs) after the optic nerve (ON) transection. METHODS: The expression and phosphorylation of c-Jun protein and apoptosis in RGCs were examined after ON transection in wild-type mice and mice in which both phosphoacceptor serines of Jun have mutated to alanines (c-Jun[AA] mice). The fluorescent tracer 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) was applied to the superior colliculi (SC), and the right ON was severed after 7 days. After two more weeks, the average number of RGCs per field was calculated. RESULTS: JNP and TUNEL-labeled apoptotic nuclei were detected in the ganglion cell layer (GCL) of the retina of the wild-type mice in response to ON transection. The numbers of TUNEL-positive nuclei in the c-Jun(AA) mice was reduced in comparison to those in wild-type mice. Retrograde labeling showed that the number of the RGCs in the retinas on the injured side of the c-Jun(AA) mice was significantly higher than in wild-type mice 14 days after the lesion. CONCLUSIONS: These results suggest that there is a partial but significant contribution of JNP to the induction of apoptosis in RGCs by ON transection.

Animals↗
PubMedDate not supplied

Urine cytology of localized primary amyloidosis of the ureter: a case report.

BACKGROUND: Morphologic findings of amyloid in urine cytology material have rarely been reported because amyloidosis of the urinary tract is a relatively uncommon disorder. We present a case of primary amyloidosis of the ureter, including catheterized urine cytologicfindings. CASE: A 78-year-old man had pollakiuria and dysuria for 5 years before admission after transurethral resection of the prostate. Clinical examination revealed left hydronephrosis and stricture of the lower part of the left ureter, and a malignant ureteral tumor was suspected clinically. In catheterized urine cytology, many clusters of epithelial cells, inflammatory cells and abundant, amorphous, waxy material were observed. The amorphous material stained light green by the Papanicolaou method and positive with direct fast scarlet (DFS), showing yellow-green birefringence under polarized light. Positivity with DFS staining was not affected by treatment with potassium permanganate. Immunocytochemically the material was AL-type amyloid protein. Atypia were absent from epithelial cells. The patient had no history of diseases that could cause secondary amyloidosis. The present case was considered to be primary amyloidosis localized to the left ureter because no particular morphologic change in the epithelial cells of the urinary tract was observed. CONCLUSION: Amyloid can be present in urine and should not be overlooked or confused with tumor diathesis when a malignant tumor is suspected clinically.

Aged↗
PubMedDate not supplied

Expression of p27(KIP1) and cell proliferation in human retina and retinoblastoma.

BACKGROUND: Retinoblastoma is a rare cancer of the eye, in which biallelic inactivation of the retinoblastoma gene is a hallmark. Although retinoblastoma protein (Rb) and p27(KIP1) block the cell cycle transition from G1- to S-phase, the interaction has not been confirmed in vivo. The aim of this study was to examine the correlation between the expression of p27(KIP1) and cell proliferation in human retina and retinoblastoma. MATERIALS AND METHODS: Human retinoblastoma, surgically removed, was fixed by 4% paraformaldehyde. Then, paraffin-embedded tissue sections were examined using immunohistochemistry with anti-p27(KIP1) and anti-proliferating cell nuclear antigen (PCNA) antibodies. RESULTS: Retinoblastoma tissue was adjacent to the normal retina in which tumor cells with homogeneous nuclei proliferated and it was impossible to identify the layer structure of the inner nuclear layer (INL) and the outer nuclear layer (ONL). In normal retina, PCNA-positive nuclei were not observed, whereas nuclear immunoreactivity for PCNA was detected in a variety of tumor cells. Many p27(KIP1)-positive nuclei were detected in INL and ONL, while p27(KIP1) immunoreactivity was not detected in retinoblastoma cells. CONCLUSION: The correlation between disappearance of p27(KIP1) and induction of proliferation activity suggests that functional loss of Rb leads to down-regulation of p27(KIP1) and uncontrolled retinal cell proliferation.

Cyclin-Dependent Kinase Inhibitor p27↗