Biomedical subjects

M Ogihara

Publications and source records attributed to M Ogihara.

At least 37 records · Page 2Linked to original sources

PubMed1998

Proliferation of adult rat hepatocytes in primary cultures induced by platelet-derived growth factor is potentiated by phenylephrine.

We investigated whether or not proliferation of adult rat hepatocytes induced by platelet-derived growth factor (PDGF) is affected by alpha1-adrenoceptor agonists such as phenylephrine during the early and late phases of primary culture. Adult rat hepatocytes underwent significant DNA synthesis after culture with 10 ng/ml of PDGF for 2 hr at a low cell density (3.3 x 10(4) cells/cm2). Under these culture conditions, the number of nuclei increased significantly during the 3.5-hr culture period. Hepatocyte DNA synthesis and proliferation induced by 10 ng/ml of PDGF decreased slightly as a result of increasing the initial plating density. An alpha1-adrenoceptor agonist, phenylephrine (10(-6) and 10(-5) M), alone did not affect hepatocyte DNA synthesis and proliferation, but markedly potentiated PDGF-induced hepatocyte DNA synthesis and proliferation. The phenylephrine effect was mimicked by phorbol myristate acetate (10(-7) M), but not by ionomycin (10(-5) M). The mitogenic effects of PDGF were almost completely blocked by treating hepatocytes with genistein (5 x 10(-6) M), U-73122 (3 x 10(-6) M), sphingosine (10(-5) M), wortmannin (10(-7) M) and rapamycin (10 ng/ml). These results demonstrate that PDGF can induce the proliferation of adult rat hepatocytes rapidly in primary culture, regardless of the initial plating density. The present results also suggest that following stimulation with PDGF, activation of tyrosine kinase, phospholipase C, phosphatidylinositol 3-kinase, protein kinase C (PKC) and p70 ribosomal protein S6 kinase is essential for the proliferation of adult rat hepatocytes. The co-mitogenic effects of phenylephrine may involve PKC activation.

Adrenergic alpha-Antagonists↗

PubMed1998

[A case of multiple sclerosis with initial symptoms of narcolepsy].

We reported a 13-year-old boy diagnosed as multiple sclerosis associated with narcolepsy. He had suffered from retrobulbar optic neuritis at the age of 11 years which was improved gradually by prednisolone. Four months later he was admitted because of excessive somnolence. The diagnosis of narcolepsy was made based on hypnagogic hallucination, sleep paralysis, changes of personality and the sleep onset sREM (SOREM). The elevation of anti-measles antibody and a positive oligoclonal band in CSF, low density areas in the bilateral internal capsule on CT scan, and high signal areas in the same region on T2-weighted MRI confirmed the diagnosis of multiple sclerosis. An abnormal arousal response occurred occasionally in sleep stage 2 and 4, which started with electrical silence followed by a period with irregular high voltage slow waves and repetitive focal spike train. Those clinical symptoms and abnormalities of biochemical and electrophysiological studies normalized after treatment with prednisolone. However, abnormalities on MRI showed no improvement even after long term administration of prednisolone (2.5 mg/day).

Adolescent↗

PubMed1998

[Management of difficult airway during induction of anesthesia in a patient with Hallermann-Streiff syndrome].

An 18-year-old female with Hallermann-Streiff syndrome underwent the fixation of prolapsus recti. She had significant microgenia, mental retardation and pharyngeal airway stenosis. During induction of anesthesia with halothane and nitrous oxide, severe upper airway obstruction and respiratory depression occurred. The mask ventilation with jaw lift maneuver was impossible. Lowering anesthetic level restored her spontaneous breathing and airway patency. Although the direct laryngoscopic view under light anesthesia with halothane was limited to the epiglottic tip, blind orotracheal intubation using stylet was accomplish after several attempts. At the end of anesthesia, the tracheal tube was extubated after the patient had become fully awake and had recovered completely from neuromuscular blockade monitored by electromyography.

Adolescent↗

PubMed1997

Proliferation of adult rat hepatocytes in primary culture induced by insulin is potentiated by cAMP-elevating agents.

We investigated whether or not insulin and cAMP-elevating agents induce the proliferation of adult rat hepatocytes during the early and late phases of primary culture. Adult rat hepatocytes synthesized a significant amount of DNA when cultured in the presence of 10(-7) M insulin for 3 h. Under these conditions, the number of nuclei increased within 4 h. Hepatocyte DNA synthesis and proliferation were not essentially affected by the initial plating densities. Other cAMP-elevating agents, such as glucagon, forskolin and dibutyryl cAMP, as well as beta-adrenoceptor agonists (i.e., metaproterenol and isoproterenol) alone had no effect on either hepatocyte DNA synthesis or proliferation in primary culture. In contrast, these agents potentiated both processes at concentrations as low as 10(-7) M when cultured in combination with 10(-7) M insulin. The stimulatory effects of beta-adrenoceptor agonists and other cAMP-elevating agents were significantly blocked by the cAMP-dependent protein kinase inhibitor, H-89 (N-[2-(p-(bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride; 10(-7) M). The mitogenic effect of insulin upon hepatocytes was almost completely suppressed by genistein (5 x 10(-6) M), wortmannin (10(-7) M) and by rapamycin (10 ng/ml). These results show that insulin rapidly induced the proliferation of adult rat hepatocytes in primary culture. The mitogenic effects of insulin were potentiated by beta-adrenoceptor agonists and cAMP-elevating agents. The effects of beta-adrenoceptor agonists and cAMP-elevating agents may be mediated through cAMP-dependent protein kinase. In addition, the activation of receptor tyrosine kinase, phosphoinositide 3-kinase and p70 ribosomal protein S6 kinase may be involved in the insulin signal transduction pathway.

1-Methyl-3-isobutylxanthine↗

PubMed1997

Density-dependent proliferation of adult rat hepatocytes in primary culture induced by epidermal growth factor is potentiated by cAMP-elevating agents.

We investigated whether or not epidermal growth factor (EGF) and cAMP-elevating agents induce the proliferation of adult rat hepatocytes during the early (4 h after adding EGF) and late phases (21 h after adding EGF) of primary cultures. Adult rat hepatocytes did not significantly proliferate after culture with 20 ng/ml EGF for 4 h at a density of 1 X 10(5) cells/cm2. In contrast, when the density was decreased by about one-third to 3.3 X 10(4) cells/cm2, the number of nuclei increased about 1.2-fold after culture with 10-20 ng/ml EGF for 4 h. Under these culture conditions, DNA synthesis began within 2-4 h of exposure to 20 ng/ml of EGF, although at the high cell density, DNA was not synthesized during this period. The beta-adrenoceptor agonists, metaproterenol and isoproterenol, and other cAMP-elevating agents, such as glucagon, forskolin, and dibutyryl cAMP, potentiated both hepatocyte DNA synthesis and proliferation about 1.4-fold when cultured in combination with 20 ng/ml EGF. The stimulatory effects of metaproterenol and other cAMP-elevating agents were specifically blocked by the cAMP-dependent protein kinase inhibitor, H-89 (10(-7) M). The effect of EGF was almost completely suppressed by genistein (5 X 10(-6) M) and rapamycin (10 ng/ml), but it was unaffected by wortmannin (10(-7) M). These results demonstrate that mature rat hepatocytes can proliferate very rapidly in low-density cultures with EGF, the effects of which were potentiated by beta-adrenoceptor agonists and cAMP-elevating agents. In addition, the activation of receptor tyrosine kinase and p70 ribosomal protein S6 kinase may be involved in EGF-induced hepatocyte DNA synthesis and proliferation.

Adrenergic beta-Agonists↗

PubMed1997

[Neoadjuvant intra-arterial chemotherapy based on chemosensitivity tests for locally invasive bladder cancer].

We investigated the clinical usefulness of individualization of chemotherapeutic regimen in neoadjuvant intra-arterial chemotherapy for locally invasive bladder cancer. Anticancer drugs were selected according to the results of an in vitro chemosensitivity test (collagen matrix assay or succinic dehydrogenase inhibition test). Nine patients with locally invasive bladder cancer received 1 to 4 courses of neoadjuvant intra-arterial chemotherapy, followed by radical cystectomy. Histopathological responses in the cystectomized specimens were grade 3 in 3 cases, grade 2 in 2, grade 1b in 2 and no response in 2. Pathologically, a complete response and downstaging were observed in 3 and 4 cases, respectively. Seven of the 9 patients were alive no evidence of disease with a mean follow-up period of 38.9 months, whereas 2 patients died of metastasis within 2 years. Six of the 7 patients who showed a complete response or down staging have been free of recurrence. These findings suggest that our chemotherapeutic strategy may improve the prognosis for locally invasive bladder cancer.

Aged↗

PubMed1997

Proliferation of adult rat hepatocytes by hepatocyte growth factor is potentiated by both phenylephrine and metaproterenol.

We investigated whether or not beta and alpha adrenergic agonists could affect proliferation of adult rat hepatocytes induced by hepatocyte growth factor (HGF) during the early and late phases of primary culture. Adult rat hepatocytes underwent significant DNA synthesis after culture with 5 ng/ml HGF for 3 h at a low cell density (3.3 x 10(4) cells/cm2). Under these culture conditions, the number of nuclei increased significantly during a subsequent 4-h culture period. Hepatocyte DNA synthesis and proliferation induced by 5 ng/ml HGF was reduced at high cell densities near confluence. A beta adrenergic agonist, metaproterenol (10(-7) M), and dibutyryl cAMP significantly potentiated hepatocyte DNA synthesis and proliferation at a concentration as low as 10(-7) M when cultured in combination with 5 ng/ml HGF. Similarly, an alpha-1 adrenergic agonist, phenylephrine (10(-6)-10(-4) M) markedly potentiated HGF-induced hepatocyte DNA synthesis and proliferation. The phenylephrine effect was mimicked by a phorbol ester (10(-6) M), but not by ionomycin (10(-6) M). The mitogenic effects of HGF were almost completely blocked by simultaneous treatment of hepatocytes with genistein (5 x 10(-6) M), U-73122 (10(-6) M), wortmannin (10(-7) M), sphingosine (3 x 10(-6) M) and rapamycin (10 ng/ml). These results demonstrate that HGF can rapidly induce proliferation of adult rat hepatocytes in primary culture. However, this effect is dependent on the initial plating density. The co-mitogenic effects of metaproterenol and phenylephrine may involve both protein kinase A and protein kinase C activation, respectively. The results also suggest that following stimulation with HGF, activation of tyrosine kinase, phosphatidylinositol 3-kinase, phospholipase C and p70 ribosomal protein S6 kinase is essential for hepatocyte proliferation.

Adrenergic beta-Antagonists↗

PubMed1997

[Anesthesia for a patient with macrothrombocytopenia].

A 56-year-old female with macrothrombocytopenia was scheduled for colectomy and hepatectomy. She had not shown significant bleeding tendency. Her preoperative platelet counts were 0.5-1.6 x 10(4) microliters-1 with the use of an automated cell counter. However, microscopic examination showed platelet number ranging 0.9-3.4 x 10(4) microliters-1 and many macrothrombocytes. Therefore, platelet biomass (platelet number x platelet volume) seemed almost normal. Bleeding time was 3 minutes and platelet function was normal. She received preoperatively high dose gamma-globulin administration and intraoperatively platelet transfusion. The operation was performed under combined epidural and general anesthesia. Intra- and post-operative course was uneventful.

Anesthesia, Epidural↗

PubMed1996

Age-related changes in proliferating cell nuclear antigen levels.

To clarify the effect of aging on rat liver regeneration, we compared proliferating cell nuclear antigen (PCNA) levels in control and regenerating livers from young and aged rats 48 h after partial hepatectomy. The nucleoplasm and cytoplasm from regenerating livers of 2-month and 24 month-old rats were fractionated by phosphocellulose column chromatography, aliquots of fractions were transferred to nitrocellulose filters and the amounts of PCNA in each fraction were measured by an immunostaining method. Two forms of PCNA, L type (eluted at low concentrations of KC1) and H type (eluted at high KC1 concentrations) were observed in the nucleoplasm from both control and regenerating young rat liver. On the other hand, the cytoplasm contained P type (eluted in the pass-through fraction), L type and H type PCNA. In control liver from aged rats, three types of PCNA in the cytoplasm and two types in the nucleoplasm were present at decreased levels. In regenerating liver from young rats, the increases in L type in the cytoplasm and H type in the nucleoplasm were remarkable. However, none of the three PCNA types increased significantly during liver regeneration in aged rats. Treatment with DNase resulted in the disappearance of the H type with a concomitant increase in the P and L types. These results suggest that the H type is a complex form consisting of the P and L types of PCNA and DNA. These results suggest that the increase in the L type in the cytoplasm reflects newly synthesized PCNA production for cellular proliferation and that the increase in the H type in the nucleoplasm is a reflection of binding to DNA and the fundamental role of PCNA itself in liver regeneration in young rats. On the other hand, there was little increase in any of the three types in regenerating liver from 24-month-old rats. Thus, PCNA content may be closely related to the decrease in the rate of cellular proliferation in aged animals.

Aging↗

PubMed1996

Comparative study of bone mineral density estimated by various methods of single- and dual-energy quantitative computed tomography: the capability of the four-equation four-unknown method.

A dual-energy (DE) quantitative computed tomography (QCT) method, the four-equation four-unknown method (DEQCT 4E-4U), was assessed and compared to single-energy (SE) QCT and standard DEQCT (two-line method). The results of this study indicate that bone mineral density (BMD) was more accurately estimated by the present method than by the SEQCT or standard DEQCT techniques on the basis of a phantom study when a large fat content was present. The results of both the phantom study and a human study also showed that the present method corrected for fat in estimating BMD in the presence of high-fat content. These findings suggest that use of this method for estimating BMD can provide useful information in studies assessing the metabolic state of bone. We propose that CT numbers estimated from excised vertebral bone marrow can serve as a soft-tissue correction for the present method.

Adult↗

PubMed1996

Analysis of simultaneous transport and metabolism of ethyl nicotinate in hairless rat skin.

PURPOSE: Simultaneous skin transport and metabolism of ethyl nicotinate (EN), a model drug, were measured and theoretically analyzed. METHODS: Several permeation studies of EN or its metabolite nicotinic acid (NA) were done on full-thickness skin or stripped skin with and without an esterase inhibitor. Permeation parameters such as partition coefficient of EN from the donor solution to the stratum corneum and diffusion coefficients of EN and NA in the stratum corneum and the viable epidermis and dermis were determined by these studies. Enzymatic parameters (Michaelis constant Km and maximum metabolism rate Vmax) were obtained from the production rate of NA from different concentrations of EN in the skin homogenate. Obtained permeation data were then analyzed by numerical method based on differential equations showing Fick's second law of diffusion in the stratum corneum and the law with Michaelis-Menten metabolism in the viable epidermis and dermis. RESULTS: Fairly good steady-state fluxes of EN and NA through the skin were obtained after a short lag time for all the concentrations of EN applied. These steady-state fluxes were not proportional to the initial donor concentration of EN: EN and NA curves were concave and convex, respectively, which suggests that metabolic saturation from EN to NA takes place in the viable skin at higher EN application. The steady-state fluxes of EN and NA calculated by the differential equations with resulting permeation and enzymatic parameters were very close to the obtained data. CONCLUSIONS: The present method is a useful tool to analyze simultaneous transport and metabolism of many drugs and prodrugs, especially those showing Michaelis-Menten type-metabolic saturation in skin.

Animals↗

PubMed1996

Cell-density-dependent expression of the alpha(2)-adrenergic response by epidermal growth factor (EGF) in primary cultures of adult rat hepatocytes.

Effects of epidermal growth factor (EGF) on the expression of alpha(2)-adrenergic responses were examined in primary cultures of adult rat hepatocytes. The alpha(2)-responses were assessed by the inhibition of the rate of forskolin-stimulated cAMP formation by the selective alpha(2)-adrenergic agonists, oxymetazoline and UK-14304. Hepatocytes cultured with EGF (20 ng/ml) at a high cell density (1.0 x 10(5)/cm2) showed almost no response to the alpha(2)-adrenergic agonists, oxymetazoline and UK-14304 (1-100 mu M). In contrast, when cultured at a low cell density (3.3 x 10(4) cells/cm2) with EGF, forskolin-stimulated cAMP production was inhibited by oxymetazoline and UK-14304 in a dose-dependent manner. The alpha(2)-response was blocked by the alpha2-antagonist yohimbine (10 mu M). It was also reversed by treatment of hepatocytes with pertussis toxin (100 ng/ml). In addition, the effects of EGF on the appearance of alpha(2)-responses were almost completely inhibited by treatment of the hepatocytes with genistein (10 mu M) or cytochalasin B (10 mu M). The alpha(2)-response was abolished when cycloheximide (5 mu M) was added to the cultures. These results demonstrate that when cultured at a low cell density with EGF, adult rat hepatocytes acquire a significant alpha(2)-adrenergic response. The expression of this alpha(2)-response is associated with de novo protein synthesis.

Adrenergic alpha-Agonists↗

PubMed1996

Cell-density-dependent expression of the beta-adrenergic response by epidermal growth factor (EGF) in primary cultures of adult rat hepatocytes.

The effects of epidermal growth factor (EGF) and cell density on the appearance of beta-adrenergic responses were examined in primary cultures of adult rat hepatocytes. The beta-adrenergic response was measured as the ability to accumulate cAMP by beta 2-agonist metaproterenol in monolayers that had been cultured without or with 20 ng/ml EGF. Hepatocytes cultured with EGF at a high cell density (1.0 x 10(5) cells/cm2) showed a relatively lower response to 10 microM metaproterenol. In contrast, when cultured at a low cell density (3.3 x 10(4) cells/cm2) with EGF, the cells showed a higher response to the beta-adrenergic agonist. These responses were blocked by the beta-adrenergic antagonist propranolol (10 microM). The beta-adrenergic response increased rapidly with culture time. The addition of cycloheximide (5 microM) to the culture abolished the expression of beta-adrenergic response. The enhanced beta-adrenergic response by 20 ng/ml EGF was partially inhibited by the addition of cytochalasin B (20 microM) to the culture. The cAMP-producing response to metaproterenol (10 microM) was dose-dependently inhibited by the specific x2-agonist UK-14304. Pretreatment of the hepatocytes with pertussis toxin (100 ng/ml) potentiated the beta-adrenergic response. These results demonstrate that augmented beta-adrenergic responsiveness can be acquired by adult rat hepatocytes cultured with 20 ng/ml EGF at a low cell density, and the beta-adrenergic response involves de novo synthesis of protein(s). The results also show that significant alpha 2- and beta-adrenergic responses coexist in the primary cultures of adult rat hepatocytes.

Adrenergic alpha-Agonists↗

PubMed1996

Difference in the enhancing effects of ultrasound on the skin permeation of polar and non-polar drugs.

The effect of ultrasound (150 kHz, 111 mW/cm2) on the permeability of isosorbide dinitrate (ISDN) and antipyrine (ANP) through excised hairless rat skin was evaluated using an Arrhenius plot. The permeability coefficients of ISDN across skin (at various temperatures) in the presence and absence of ultrasound were virtually isolinear on the Arrhenius plot. It has been suggested that the temporal increase in the ISDN flux, which was observed when ultrasound was applied in our previous study, was only a result of the thermal effect of ultrasound, i.e., an increase in the temperature of the donor solution. On the other hand, ultrasound influenced the Arrhenius plot of ANP, suggesting that the enhancement effect for ANP permeation could be not explained only by the thermal effect of ultrasound. In addition, the effective diffusion (D) and partition coefficients (K) of ISDN and ANP were estimated using their skin permeation profiles across the ultrasonic pretreated skin. The coefficients of ISDN with ultrasonic pretreatment were comparable to those without pretreatment. On the other hand, the D value of ANP with ultrasonic pretreatment was increased about 4 times by ultrasonic pretreatment, in spite of an insignificant change in the K value. These results suggest that the ultrasound used in the present study increased the effective diffusivity across the aqueous region in the stratum corneum to enhance the skin permeation of the polar compound, ANP.

Animals↗

PubMed1996

[Sociomedical study of centenarians in Nagoya City].

The purpose of this study was to assess the background to the longevity of 36 centenarians in Nagoya city and to compare 14 institutionalized centenarians out of those 36 with 202 individuals in the 70-99 age group in our special nursing home, particularly with regard to blood chemistry and immunity tests. The reasons for their social longevity in terms of profile appeared to be attention to eating habits, abstention from smoking and drinking, occupations with adequate exercise. The incidence of dementia was 65.6% among them. We evaluated centenarians from the viewpoint of Hasegawa's Dementia Scale (HDS) and comprehensive functional assessment of the elderly consisted of the revised version of Hasegawa's Dementia Scale (HDSR), and examination of activity of daily living (ADL), physical perception, and social life. All were positively associated. Thus centenarians independent of physical assistance demonstrated significantly higher systolic blood pressure, and respective scores for HDS, HDSR, ADL, physical perception and social life than their dependent counterparts, were less likely to be institutionalized and suffered from fewer disorders. In particular none were diagnosed as positive for cerebral hemorrhage, infarction and dementia. Age demonstrated significant positive or negative correlation with the following values in blood chemistry and immunity tests: blood cell counts, hemoglobin concentration, hematocrit value, albumin, total protein, total cholesterol, low density lipoprotein cholesterol, creatinine, blood cell nitrogen, uric acid, helper T cell, and IgA immunoglobulin.

Activities of Daily Living↗

PubMed1996

[Efficacy of the chemosensitivity test using collagen gel matrix-supported culture system for urogenital tumors].

BACKGROUND: We evaluated the usefulness of in vitro tumor culture system using a specialized collagen gel matrix (CGM assay) as a chemosensitivity test. PATIENTS AND METHODS: Chemosensitivity results of CGM assay were compared with other in vivo and in vitro assays on an implantable murine bladder tumor cell line (MBT-2). In addition we investigated the possibility of the clinical use of CGM assay using clinical specimens obtained from urogenital malignant tumor patients by comparing the result with that of the other chemosensitivity test, SDI testing using single cells (conventional SDI test). Methods of CGM assay were as follows. Tumor tissues on the collagen gel matrices were incubated under the existence of anticancer drugs following 4 days preculture. Drug sensitivities were evaluated by counting the number of viable cells adjusted to the tumor weight. RESULTS: Inhibition rates in MBT-2 were high in the order of mitomycin C, cisdiamminedichloroplatinum (II), (2"R)-4'-0-tetrahydropyranyl adriamycin. Four of 6 anticancer drugs were decided as chemosensitive drugs. These results corresponded to the results of the antitumor effects on subcutaneously transplanted MBT-2 in vivo, moreover was correlated well with those of the conventional SDI test. Twenty of 22 cases, including 11 of 13 bladder cancer cases, 1 of 3 renal cancer cases, 2 of 3 testicular cancer cases and 1 of 1 adrenal cortical cancer cases, were evaluable in the clinical study of the CGM assay. Corresponding rates between the results of the CGM assay and those of the conventional SDI test performed simultaneously in 12 cases were excellent for each anticancer drug. CONCLUSION: This CGM assay can serve as an effective tool for chemosensitivity testing because of its convenience and high evaluable rate.

Animals↗

PubMed1996

Combination therapy of Pseudomonas aeruginosa pyelonephritis in neutropenic mice with human antilipopolysaccharide monoclonal antibody and cefsulodin.

PURPOSE: These studies were designed to determine the combined inhibitory effect of a human monoclonal antibody (MAb) and cefsulodin on Pseudomonas aeruginosa renal infection in a neutropenic condition. MATERIALS AND METHODS: Protection against the infection of mice was estimated by survival rate and bacterial numbers in the kidney and blood. Opsonophagocytic assay by human polymorphonuclear neutrophils (PMNs) and fluorescence activated cell sorter (FACS) analysis were also examined. RESULTS: Treatment of infected mice with MAb combined with a suboptimal dose of cefsulodin prevented the mice from developing pyelonephritis and bacteremia and resulted in a significantly higher survival rate than treatment with either MAb or cefsulodin alone (p < 0.01). When bacteria were preexposed to cefsulodin, a significant enhancement in opsonophagocytic killing with MAb was observed. Fluorescence activated cell sorter analysis suggested that the bacteria incubated with 1/4 minimal inhibitory concentration (MIC) of cefsulodin showed greater binding of MAb to bacteria than the control. CONCLUSION: The combination therapy with human antilipopolysaccharide MAb and cefsulodin is useful for P. aeruginosa pyelonephritis in neutropenic hosts.

Animals↗

PubMed1995

Stimulation of DNA polymerase gamma activity by proliferating cell nuclear antigen.

DNA polymerase was partially purified from mitochondrial extracts of rat liver by phosphocellulose, DEAE-cellulose, heparin-Sepharose CL-6B and DNA-agarose column chromatography. By these purification steps, DNA polymerase and proliferating cell nuclear antigen (PCNA) were completely separated at the step of heparin-Sepharose CL-6B column chromatography. The isolated DNA polymerase was inhibited by ddTTP, but not by aphidicolin. The enzyme sedimented at about 8 S on 5-20% analytical sucrose density gradient centrifugation. These data showed that the DNA polymerase isolated from mitochondria is gamma in type. After the separation of DNA polymerase gamma and PCNA, the two fractions were remixed and DNA polymerase gamma activity was measured. DNA polymerase gamma activity was stimulated about three-fold or more in the presence of the PCNA fraction. This stimulation was inhibited by the addition of anti-PCNA rabbit IgG2a. In addition, highly purified human recombinant PCNA stimulated the DNA polymerase gamma activity. These results indicate that DNA polymerase gamma, like DNA polymerase delta, is activated by PCNA.

Animals↗