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Biomedical subjects

N Shinohara

Publications and source records attributed to N Shinohara.

At least 73 records · Page 4Linked to original sources
PubMed1997

Co-receptor-independent signal transduction in a mismatched CD8+ major histocompatibility complex class II-specific allogeneic cytotoxic T lymphocyte.

The contribution of co-receptors in signal transduction upon T cell receptor (TCR)-mediated recognition of major histocompatibility complex (MHC) class II antigen by mature T lymphocytes expressing TCR derived from the apparently co-receptor-independent, I-Ak-specific allogeneic CD8+ CTL clone QM11 has been examined. Mature double-negative, CD8+ and CD4+ bulk T cell lines and clones expressing TCR(QM11) were developed from TCR(QM11) transgenic mice. All these T cells, irrespective of co-receptor expression, showed specific lytic activity on cells expressing I-Ak. Furthermore, co-receptorless mutants were obtained from a CD4+ and CD8+ clone. The responses of these co-receptorless mutants upon specific recognition of the alloantigen, as judged by cytolytic activity, granule exocytosis, lymphokine production, proliferation, and tyrosine phosphorylation of the zeta chain, were comparable to those of the original clones. Thus, the results proved the co-receptor independence of the recognition of I-Ak by TCR(QM11) and further indicated there is no indispensable unique signal transduced by co-receptors. However, when the amount of the available antigen was limited by anti-I-Ak antibody, the CD4+ T cell clone showed a remarkable resistance to the inhibition whereas the mismatched CD8+ clone was readily inhibitable. The anti-I-Ak-resistant component of the CD4+ clone showed dependency on the CD4 molecule. Taken collectively, the results indicate that the role played by a co-receptor molecule in mature T cells is purely quantitative amplification of the signal through the formation of a TCR/MHC/co-receptor ternary complex, and also indicate that the role of co-receptor molecules as TCR-independent adhesion molecules is at best minimal.

Animals↗
PubMed1997

Reduced tissue oxygenation and its reversibility by glycemic control in diabetic patients.

Relationship of transcutaneous oxygen pressure (TcP(O2)) to glycemic control and diabetic complications was investigated in patients with non-insulin dependent diabetes mellitus. TcP(O2) was measured in 103 patients with non-insulin dependent diabetes mellitus. Correlation of TcP(O2) to HbA1c, fasting blood sugar (FBS), age, duration of diabetes, serum lipids, hypertension, and diabetic complications were examined. We divided the patients into three groups according to their glycemic control: good control group (HbA1c < 7.0%), fair control group (HbA1c, 7.0-8.9%) and poor control group (HbA1c > or = 9.0). We compared TcP(O2) of these three groups with 19 non-diabetic controls. In 103 patients, TcP(O2) at baseline correlated with HbA1c, FBS and age (P < 0.001, P < 0.01 and P < 0.05, respectively), but did not correlate with duration of diabetes mellitus, neuropathy, nephropathy or retinopathy. TcP(O2) of good and fair control group was not reduced comparing to the non-diabetic control (63 +/- 11, 59 +/- 10 and 64 +/- 12 mmHg, respectively). The poor control group had significantly reduced TcP(O2) (55 +/- 10 mmHg) comparing to non-diabetic control (P < 0.005) and good control group (P < 0.005). Furthermore, in an independent study, TcP(O2), arterial oxygen pressure (Pa(O2)), oxygen pressure of dorsal pedal vein (PV(O2)) and erythrocyte 2,3-diphosphoglycerate (2,3-DPG) in eight patients with poor glycemic control were followed prospectively. Six patients with improvement of glycemic control showed a significant increase of TcP(O2) and Pa(O2) (P < 0.001 and P < 0.005, respectively). However, two patients without improvement of hyperglycemia had no change in TcP(O2) and Pa(O2). PV(O2) and 2,3-DPG levels of erythrocytes were not changed in six patients. These findings suggest that tissue oxygenation in diabetic patients was deteriorated in relation to hyperglycemia and was reversed with glycemic control. Improvement of Pa(O2) might contribute partly to the increase of TcP(O2).

2,3-Diphosphoglycerate↗
PubMed1997

The significance of Ras guanine nucleotide exchange factor, son of sevenless protein, in renal cell carcinoma cell lines.

PURPOSE: The aim of the present study is to clarify the significance of the Ras guanine-nucleotide exchange reaction in the proliferation of human renal cell carcinoma cell lines. MATERIALS AND METHODS: We examined the expression of human son of sevenless-1 (hSos-1) protein and the epidermal growth factor (EGF) receptor in human renal cell carcinoma cell lines by Western blot analysis. Additionally, a dominant negative H-ras mutant, N116Y, which is known to inhibit the Ras guanine-nucleotide exchange reaction, was transfected into these cell lines by lipofection. RESULTS: Human renal cell carcinoma cell lines expressed much higher amounts of the EGF receptor and hSos-1 protein than normal kidney tissue. Moreover, the N116Y ras mutant could strongly suppress cellular proliferation in these cell lines. CONCLUSIONS: Augmentation of the Ras guanine-nucleotide exchange reaction might be essential to the proliferation of human renal cell carcinoma cells.

Carcinoma, Renal Cell↗
PubMed1997

Three novel mutations and a de novo deletion mutation of the DAX-1 gene in patients with X-linked adrenal hypoplasia congenita.

The DAX-1 [DSS (dosage sensitive sex)-AHC critical region on the X, gene 1] gene is responsible for X-linked adrenal hypoplasia congenita (AHC). However, DAX-1 protein structure-function relationships are not well understood. Identification of missense mutations may help to reveal these relationships. We analyzed the DAX-1 gene from seven patients in six kindreds with X-linked AHC and identified one frameshift mutation, two missense mutations, and three deletion mutations. Case 1 had a 388delAG frameshift mutation, inducing a premature stop codon at position 70. Case 2 had a missense mutation, Lys382Asn, which encodes an asparagine (Asn) for lysine (Lys) at position 382. Sibling cases of 3-1 and 3-2 had a missense mutation of Trp291 Cys, which encodes a substitution of cysteine (Cys) for tryptophan (Try) at position 291. The tryptophan (Trp) at position 291 and lysine (Lys) at position 382 in human DAX-1 protein are highly conserved among other related orphan nuclear receptor superfamily members. Cases 4, 5, and 6 showed deletion mutation. In case 6, a de novo deletion mutation was revealed by both southern hybridization and polymerase chain reaction (PCR) of a GGAA tetranucleotide tandem repeat. These findings suggest that: 1) Trp at position 291 and Lys at position 382, located in the C-terminal presumptive ligand binding domain, are important to the functional role of the DAX-1 protein in adrenal embryogenesis and/or in hypothalamic-pituitary activity; and 2) molecular analysis of the DAX-1 gene may help genetic counseling, even in cases with deletion mutation, because a detection of de novo deletion may exclude another affected or carrier child.

Adrenal Insufficiency↗
PubMed1997

Cytokeratin subtyping in chordomas and the fetal notochord: an immunohistochemical analysis of aberrant expression.

Chordoma is a well-known bone tumor that shows epithelioid features and in which the expression of cytokeratins (CKs) has been reported to appear very frequently. Numerous immunohistochemical analyses of CK expression have been conducted using such monoclonal antibodies as CAM5.2, which react with CK8, CK18, and CK19, and AE1/AE3, which react with CKs 1-8, 10, 14-16 and 19 in chordoma. No detailed analysis, however, of the expression of each component of CK has yet been conducted in chordoma; thus, the subsets of CK expressed there have yet to be clarified. With the use of immunohistochemical techniques with a panel of monoclonal antibodies against each subset of CK, the authors studied the expression of CKs in 16 specimens of classic chordoma and 14 specimens of the fetal notochord to clarify the subsets of CK expressed in chordoma and to evaluate the similarities and differences of CK expression between chordoma and the fetal notochord. All of the chordoma specimens showed a strong positive immunoreactivity for CK8 and CK19, whereas nine (56.3%) chordoma specimens showed a positive immunoreaction for CK18. In addition, four chordoma specimens were focally positive for keratin-903, which reacts with high molecular weight CKs such as CK1, CK5, CK10, and CK14; one specimen also showed a strong CK7 expression. All of the notochord specimens were also positive for CK8 and CK19, but none showed a positive immunoreaction for keratin-903, CK7, or CK18. In addition, none of the chordoma or notochord specimens showed immunoreactivity for CK20. The expression of CK8 and CK19, observed in all of the chordoma and notochord specimens, was thus considered to be maintained throughout the neoplastic transformation, although some aberrant CK expressions (CK7, CK18, and keratin-903) also occurred in the chordoma specimens examined in this study.

Adult↗
PubMed1997

Expression of c-met proto-oncogene product (c-MET) in benign and malignant bone tumors.

The expression of c-met proto-oncogene product (c-MET) has been reported to be related to invasive growth or tumor stage in some tumors, but little is known concerning the significance of c-MET expression in bone tumors. With use of formalin-fixed, paraffin-embedded tissue specimens and polyclonal antibody for c-MET, we studied the expression of c-MET in 122 cases of malignant bone tumors (43 osteosarcomas, 24 chondrosarcomas, 21 malignant fibrous histiocytomas of bone, 16 Ewing's sarcoma versus primitive neuroectodermal tumors, 18 chordomas), 65 cases of benign tumors and tumor-like lesions (including 8 giant cell tumors of bone, 8 chondroblastomas, 12 enchondromas, 7 osteochondromas, 10 fibrous dysplasias), 7 cases of articular cartilaginous tissue, and 10 cases of fetal vertebral tissue consisting of foci of enchondral ossification and notochordal tissue. In malignant tumors, c-MET expression was most frequently detected in chordoma (94.4%), followed by chondrosarcoma (54.2%) and osteosarcoma (23.3%). Among the osteosarcoma specimens, c-MET expression was frequently detected in the chondroblastic subtype (66.7%), but the incidence was low in the cases with other subtypes of osteosarcoma. We found no significant correlation between the c-MET expression and the histologic grade of malignancy in either osteosarcoma or chondrosarcoma. c-MET expression was either rarely observed or completely negative in malignant fibrous histiocytomas of bone (4.8%) and primitive neuroectodermal tumors (0%). In benign tumors and tumor-like lesions, c-MET expression was frequently detected in cartilaginous tumors, such as chondroblastoma (62.5%), enchondroma (66.7%), and osteochondroma (71.4%), but no expression was observed in giant cell tumors of bone or any other benign tumors or tumor-like lesions. In normal tissue, c-MET expression was frequently detected in the articular cartilage (100%) and notochord (70.0%) specimens examined. We conclude that c-MET expression as frequent as that observed in the notochordal tissue, chordomas, articular cartilage, and cartilaginous tumors is related to the development of both normal tissue and chondroid tumors.

Adolescent↗
PubMed1996

Concanamycin A, a powerful tool for characterization and estimation of contribution of perforin- and Fas-based lytic pathways in cell-mediated cytotoxicity.

Perforin- and Fas-based cytolytic pathways are two major mechanisms of cell-mediated cytotoxicity. Recently, we have shown that an inhibitor of vacuolar type H+-ATPase, concanamycin A (CMA), inhibits perforin-based cytotoxic activity, mostly due to accelerated degradation of perforin by an increase in the pH of lytic granules. Here we show that CMA failed to inhibit the cytolytic activity of CD4+ CTL clone and perforin-deficient CD8+ CTL clone, which exclusively mediate Fas-based cytotoxicity, although CMA inhibited acidification and induced drastic vacuolation of cytoplasmic granules in these clones. In a wide range of alloantigen-specific CTL, a significant amount of the lysis of Con A blasts from normal mice and of Fas-positive tumor cells remained unaffected even in excess concentrations of CMA. However, CMA almost completely inhibited the lysis of Con A blasts from lpr mice and of Fas low expressing or negative tumor cells. Cytolysis by alloantigen-specific CD8+ CTL derived from gld mice was completely prevented by CMA. Furthermore, CMA-insensitive cytolysis exerted by CD8+ CTL clone was completely inhibitable by soluble Fas molecules. Thus, these data clearly indicate not only that CMA-insensitive cytolysis mediated by alloantigen-specific CTL is Fas dependent, but also that CMA is a selective inhibitor to block only the perforin-based killing pathway. In contrast, brefeldin A blocked the Fas-based cytotoxicity, but only marginally reduced the perforin-based cytotoxicity. Moreover, CMA and brefeldin A in combination completely abrogated all cytolytic activity of alloantigen-specific CTL. Taken together, these results reveal that CTL mainly exert perforin-based cytotoxicity and complementary Fas-based cytotoxicity, and that CMA is a powerful tool to clarify the contributions of the two distinct cytolytic pathways.

Animals↗
PubMed1996

The proportion of free to total prostate specific antigen: a method of detecting prostate carcinoma.

BACKGROUND: Prostate specific antigen (PSA) is the most useful marker for prostate carcinoma (CaP). However, the sensitivity and specificity for PSA are not sufficient for the diagnosis of organ-confined prostate carcinoma. Recent studies have revealed that anti-PSA antibody identifies both PSA complexed to alpha-1-antichymotrypsin and free PSA, whereas anti-gamma-seminoprotein antibody recognizes free PSA exclusively. To enhance the ability of PSA to detect CaP in patients with total PSA levels of 10 ng/mL or lower, we developed the ratio of gamma-seminoprotein and PSA (free/total PSA index). METHODS: We measured free/total PSA indices for 285 patients who had serum PSA levels of 10 ng/mL or lower and who were diagnosed pathologically. RESULTS: Of the 285 patients, 228 had no prostate carcinoma (NC) and 57 had CaP. The mean total PSA level for CaP (5.137 +/- 2.483 ng/mL; mean +/- standard deviation) was significantly greater (P < 0.0001) than that for NC (3.251 +/- 2.129). The mean free/total PSA index for CaP (0.774 +/- 0.468 was significantly lower (P < 0.0001) than that for NC (1.563 +/- 0.938). The sensitivity for the free/total PSA index was similar to that for total PSA (78.9% vs. 75.4%). However, the specificity, positive predictive value, and overall accuracy for the free/total PSA index (75.9%, 45%, and 76.5%, respectively) increased by 15-20% compared with those for total PSA (56.6%, 30.3%, and 60.4%, respectively). CONCLUSIONS: The free/total PSA index improved the specificity of PSA without impairing the sensitivity in detecting CaP among patients with serum PSA levels of 10 ng/mL or lower.

Humans↗
PubMed1996

Derivation of T-cell receptor alpha-chain double expresser lines from normal murine mature T cells.

Because the T-cell receptor (TCR) alpha-chain locus is known to lack allelic exclusion of rearrangements, and as a recent report revealed the existence of alpha-chain double expressers among normal human peripheral blood lymphocytes (PBL), the possible existence of TCR alpha-chain double expressers among mature murine T cells was examined. Although two-colour staining analysis of normal T-cell populations did not immediately reveal recognizable clusters of V alpha double expressers, alternative in vitro stimulations of normal murine T cells with antibodies to two different TCR V alpha chains reproducibly induced TCR alpha-chain double-expresser lines. TCR complexes with different alpha-chains on such T cells were both shown to be functional. The cell lines were heterogeneous with respect to V beta usage and the ratio of the expressed amounts of the two alpha-chains on the surface. The ratio of the two expressed alpha-chains was found to be very stable over a long period of time. These results are consistent with the earlier report on alpha-chain double expressers among human T cells and also show normal occurrence of TCR alpha-chain double expressers in murine T-cell populations.

Animals↗
PubMed1996

Vertebral growth disturbance in rapidly growing rats during 14 days of spaceflight.

The effect of 14 days of spaceflight on the vertebrae of rapidly growing rats was studied. The hardness of the vertebrae was measured with a Knoop microhardness tester, and bone mineral density was measured from X-ray photographs. Histomorphometric examination was performed with a microcomputer-aided system. No significant difference (P > 0.05) was observed between flight rats and ground controls with regard to mechanical hardness or bone mineral density. However, histological examination revealed irregular thickening of the endosteal surface of cortical bone in the flight rats, whereas it was uniform in the ground controls. The relative area of lamellar bone showed a significant reduction (P < 0.001) in the flight rats. These findings suggest that the structural disturbances were due to retardation of endosteal modeling and remodeling. We conclude that delay of vertebral maturation can occur in rapidly growing rats after even short-term exposure to microgravity.

Animals↗
PubMed1996

Truncation at the C-terminus of the DAX-1 protein impairs its biological actions in patients with X-linked adrenal hypoplasia congenita.

The DAX-1 [DSS (dosage-sensitive sex)-AHC critical region in the X, gene 1] gene has been reported to be responsible for X-linked adrenal hypoplasia congenita (AHC) and hypogonadotropic hypogonadism. However, the function and structure of the DAX-1 protein have not been characterized. In this study, molecular analysis of the DAX-1 gene from 6 patients with AHC, including 2 siblings, identified 5 novel mutations with 3 nonsense mutations and 2 frameshift mutations. Case 1 had a nonsense mutation at position 395 (Q395X). Cases 2 and 3, who were siblings, had a nonsense mutation at position 91 (Y91X). Case 4 had a 2-base deletion (AT) at nucleotides 1610 and 1611 and a 1-base insertion (G) resulting in a premature stop codon at position 462 (1610-1611 del AT ins G). Case 5 had a nonsense mutation at position 271 (Y271X). Case 6 had a 1-base deletion (C) at nucleotide 1169, which induced a frame shift and a premature stop codon at position 371 (1169 del C). All mutated DAX-1 proteins had truncated C-terminal domains. In addition, reverse transcription-PCR and direct sequencing characterized the mutant messenger ribonucleic acid in testis from case 1. Our results suggest that these 5 novel mutations are responsible for X-linked AHC and that the C-terminus of the DAX-1 protein, especially the terminal 11 amino acids, is necessary for normal adrenal cortical embryogenesis.

Adolescent↗
PubMed1996

Sequential occurrence of hepatocellular carcinoma following Waldenström's macroglobulinemia: the pathogenetic role of chronic liver disease.

We report a case of sequential occurrence of hepatocellular carcinoma (HCC) following Waldenström's macroglobulinemia (WM) in the course of chronic liver disease. The diagnosis of WM was made in a 55-year-old man with a history of alcohol abuse. He developed HCC in the course of WM, and hepatitis C virus (HCV) infection was detected by enzyme-linked immunosorbent assay. Recent reports suggest that persistent HCV infection is related to the pathogenesis of B cell malignancy. Chronic liver disease may be associated with the pathogenesis of the double malignancies in the present case.

Carcinoma, Hepatocellular↗
PubMed1996

[Treatment of advanced testis cancer with a high dose chemotherapy regimen (PVeBV)].

BACKGROUND: The prognosis of patients with testis cancer classified as being in the advanced extent according to the Indiana University staging system is still poor even when treated with cisplatin based chemotherapy. METHODS: Attempting to increase the efficacy of chemotherapy in this high risk group, we have adopted PVeBV chemotherapy (high dose CDDP+VBL+VP-16+BLM) for recent 8 patients with such advanced conditions. In this study, we analized the treatment outcome of those patients retrospectively. RESULTS: Two patients died during the first course of PVeBV chemotherapy due to cancer progression, while 6 patients treated with 3 to 4 cycles of PVeBv were eligible and assessable for response, survival, and toxicity. Five of those 6 achieved pathological CR (pCR) following surgical resection of residual masses after 3 cycles of PVeBV. The other case was saved by salvage chemotherapy with autologous BMT. All 6 patients were long-term disease free survivors in median follow up of 46 months. With the rG-CSF application and vigorous hydration, acute phase toxic effects (myelosuppression, pulmonary fibrosis and nephrotoxicity) were manageable in this intensive regimen. Long term toxic effects such as peripheral neuropathy and ototoxicity were also tolerable and quality of life in such advanced cancer patients was preserved well. CONCLUSION: To improve a cure rate of high risk testis cancer, the dose escalation of induction chemotherapy should be considered.

Adolescent↗
PubMed1996

[Prognostic factors in patients with advanced prostatic cancer (stage D2) initially treated with endocrine therapy].

BACKGROUND: In order to make clear the prognostic factors in patients with advanced prostatic cancer (stage D2) whose initial treatment was hormone therapy, we reviewed 34 men with newly diagnosed advanced prostatic cancer for 6 years from May 1987 at the Hospital of Hokkaido University. METHODS: All patients had histologically proven prostatic adenocarcinoma. The mean age was 67.9 years (range 54 to 86 years). Median follow-up period was 26 months (range 5 to 66 months). The parameters studied were age, performance status, histological grade, extent of disease on bone scan, pretreatment values of PSA, gamma-Sm, PAP and PSA/gamma-Sm ratio, posttreatment values of PSA, gamma-Sm and PAP at 6th month, and the ratio of PSA at 3rd month to PSA at 6th month. RESULTS: Univariate analysis revealed that performance status (PSO vs. PS2, p = 0.006, PS1 vs. PS2, p = 0.016) and extent of disease on bone scan (p = 0.004) in background factors, gamma-Sm (p = 0.005) among pretreatment values of markers, PAP at 6th month (p < 0.001) and the ratio of PSA at 3rd month to PSA at 6th month (p < 0.001) among posttreatment values of markers, significantly related with the cause-specific survival. Based on multivariable analysis, PAP at 6th month and the ratio of PSA at 3rd month to PSA at 6th month were significantly correlated with the cause-specific survival at 24th month and 36th month, respectively. CONCLUSION: We conclude that posttreatment values of markers are more important prognostic factors in patients with advanced prostatic cancer (stage D2) following hormone therapy than pretreatment background factors and values of markers.

Adenocarcinoma↗
PubMed1996

[Tumor size and DNA ploidy changes in renal cell carcinomas--flow cytometric analysis of DNA content in renal cell carcinomas associated with von Hippel-Lindau disease].

BACKGROUND: Renal cell carcinomas (RCCs) develop in 8-63% of von Hippel-Lindau disease (VHL) patients, and loss of 3p segments, chromosome aberrations found in 90% of sporadic RCCs, has also been observed in RCCs associated with VHL. In fact, comparative analysis showed that the chromosome aberrations in RCCs associated with VHL are similar to those found in sporadic RCCs. VHL patients have the whole spectrum of tumors from small early lesions to large ones in the same kidney, providing a unique opportunity to analyze tumors in different stages of development. Subsequently deoxyribonucleic acid (DNA) content in RCCs of VHL patients was examined and correlated to their tumor size to gain some insight in the progression of sporadic RCCs. METHODS: From 1988 to 1991, we have experienced 6 cases of RCCs associated with VHL who underwent partial or radical nephrectomy. A total number of 52 paraffin-embedded samples from 33 RCCs from 6 patients with VHL was analyzed by flow cytometry. RESULTS: The sizes of tumors ranged from 0.2 to 8.2 cm. DNA aneuploid patterns demonstrated in none of 9 tumors less than 1.6 cm, 4 of 14 tumors (29%) as large as 1.6 to 2.5 cm, and 5 of 10 tumors (50%) larger than 2.5 cm (p < 0.05). Twelve tumors less than 1.8 cm showed DNA diploid, so the smallest size of aneuploid tumors was 1.8 cm. CONCLUSION: These data suggest that DNA ploidy change (diploid to aneuploid) in RCCs probably takes place as tumors grow approximately 1.8 cm in size.

Adult↗
PubMed1996

[Effective administration of recombinant granulocyte colony-stimulating factor to prevent granulocytopenia due to combination chemotherapy for testicular germ cell tumors].

A prospective randomized study on the administration of recombinant granulocyte colony stimulating factor (rG-CSF) was conducted on 15 patients with testicular germ cell tumors. The clinical stagings of all patients except one were minimal to moderate extent according to the Indiana University staging system. Combination chemotherapy using bleomycin, etoposide and cisplatinum (BEP) was performed as the initial treatment on the eligible patients. rG-CSF was administered by two different methods; 1) routine administration on the 6th day after BEP chemotherapy (group A), and 2) the same method, but after granulocytopenia of 1,500/mm3 had developed (group B). The administration of rG-CSF in group A significantly reduced the severity of leucocytopenia and also the incidence of stomatitis compared with group B. Although rG-CSF produced no significant side effects, the thrombocytopenia was prominent in the group A patients (not significant). BEP chemotherapy itself is an easily-tolerable and well established method for treating young adult patients. The method used in group B seems to be suitable in situations where thrombocytopenia and cost effectiveness.

Adult↗
PubMed1996

[Purification of EcoO44I restriction endonuclease in Escherichia coli O44 isolated from an affected human].

A restriction endonuclases (ENase) designated EcoO44I was purified without non specific nucleases from enteropathogenic Escherichia coli O44 Hiromi strain of affected human origin. The yield was 1, 100 units/g of wet cells. The EcoO44I ENase recognized and cleaved the specific sequence of 5'-GGTCTC-3' (1/5) as was the case with Eco31I or BsaI ENase. Because of the stability and high yield, EcoO44I would be useful for recombinant DNA technology after isolation of EcoO44-positive, avirulent mutant strains of E. coli O44 Hiromi.

DNA Restriction Enzymes↗