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PubMed1999

Differentiation of adipose stromal cells: the roles of glucocorticoids and 11beta-hydroxysteroid dehydrogenase.

Glucocorticoids play an important role in determining adipose tissue distribution and function, with glucocorticoid excess states such as Cushing's syndrome resulting in central obesity. We have investigated the functional significance of local generation of cortisol within adipose tissue from inactive cortisone through the activity of the NADP(H)-dependent enzyme, 11beta-hydroxysteroid dehydrogenase type 1 (11betaHSD1). In primary cultures of paired omental (om) and sc human adipose stromal cells (ASC; n = 34), 11betaHSD1 oxo-reductase activity was significantly higher in om ASC (median, 40.2 pmol/mg protein x h; 95% confidence interval, 1.8-105) compared with sc ASC (median, 11.4; 95% confidence interval, 0-48.1; P<0.001) despite similar endogenous NADPH/NADP concentrations. Both cortisol and insulin increased the differentiation of ASC to adipocytes (as assessed by glycerol-3-phosphate dehydrogenase expression), but only cortisol increased 11betaHSD1 activity and messenger RNA levels in a dose-dependent fashion. Cortisone (500 nM) was as effective as 500 nM cortisol in inducing ASC differentiation, but this stimulatory effect was inhibited by coincubation with the 11betaHSD1 inhibitor, glycyrrhetinic acid. The higher local conversion of cortisone to active cortisol through expression of 11betaHSD1 in om compared with sc ASC may explain the specific action of glucocorticoids on different adipose tissue depots. 11betaHSD1 expression in om ASC is regulated at a transcriptional level and is increased by glucocorticoids, but is not entirely dependent upon ASC differentiation. Inhibition of 11betaHSD1 within om ASC inhibits cortisone-induced ASC differentiation. These findings indicate that local metabolism of glucocorticoid may control differentiation of adipose tissue in a site-specific fashion. Specific inhibitors of 11betaHSD1 may offer a novel approach for the treatment of patients with central obesity.

11-beta-Hydroxysteroid Dehydrogenases↗

PubMed2005

Characterization of activating signal cointegrator-2 as a novel transcriptional coactivator of the xenobiotic nuclear receptor constitutive androstane receptor.

Activating signal cointegrator-2 (ASC-2) is a recently isolated transcriptional coactivator protein for a variety of different transcription factors, including many members of the nuclear receptor superfamily. In this report, we demonstrate that ASC-2 also serves as a coactivator of the xenobiotic nuclear receptor constitutive androstane receptor (CAR). First, transcriptional activation by CAR was enhanced by cotransfected ASC-2 in CV-1 and HeLa cells. In contrast, CAR transactivation was significantly impaired in HepG2 cells stably expressing specific small interfering RNA directed against ASC-2. Consistent with these results, chromatin immunoprecipitation experiments revealed that ASC-2 is recruited to the known CAR target genes in a ligand-dependent manner. Secondly, CAR specifically interacted with the first LXXLL motif of ASC-2, and these interactions were stimulated by CAR agonist 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene and repressed by CAR inverse agonist androstanol, suggesting that this motif may mediate the interactions of ASC-2 and CAR in vivo. In support of this idea, DN1, a fragment of ASC-2 encompassing the first LXXLL motif, suppressed CAR transactivation, and coexpressed ASC-2 but not other LXXLL-type coactivators such as thyroid hormone receptor-associated protein 220 reversed this repression. Finally, CAR was recently found to play a pivotal role in effecting the severe acetaminophen-induced liver damage. Interestingly, transgenic mice expressing DN1 were resistant to the acetaminophen-induced hepatotoxicity and expression of a series of the known CAR target genes was specifically repressed in these transgenic mice. Taken together, these results strongly suggest that ASC-2 is a bona fide coactivator of the xenobiotic nuclear receptor CAR and mediate the specific xenobiotic response by CAR in vivo.

Acetaminophen↗

PubMed2005

Stem cell therapy for neurologic disorders: therapeutic potential of adipose-derived stem cells.

There is growing evidence to suggest that reservoirs of stem cells may reside in several types of adult tissue. These cells may retain the potential to transdifferentiate from one phenotype to another, presenting exciting possibilities for cellular therapies. Recent discoveries in the area of neural differentiation are particularly exciting given the limited capacity of neural tissue for intrinsic repair and regeneration. Adult adipose tissue is a rich source of mesenchymal stem cells, providing an abundant and accessible source of adult stem cells. These cells have been termed adipose derived stem cells (ASC). The characterization of these ASCs has defined a population similar to marrow-derived and skeletal muscle-derived stem cells. The success seen in differentiating ASC into various mesenchymal lineages has generated interest in using ASC for neuronal differentiation. Initial in vitro studies characterized the morphology and protein expression of ASC after exposure to neural induction agents. Additional in vitro data suggests the possibility that ASCs are capable of neuronal activity. Progress in the in vitro characterization of ASCs has led to in vivo modeling to determine the survival, migration, and engraftment of transplanted ASCs. While work to define the mechanisms behind the transdifferentiation of ASCs continues, their application to neurological diseases and injuries should also progress. The subject of this review is the capacity of adipose derived stem cells (ASC) for neural transdifferentiation and their application to the treatment of various neurologic disorders.

Adipocytes↗

PubMed2000

Does ascorbate in the mesophyll cell walls form the first line of defence against ozone? Testing the concept using broad bean (Vicia faba L.).

Broad bean (Vicia faba L.) plants were exposed, in duplicate controlled environment chambers, to charcoal/Purafil-filtered air (CFA-grown plants) or to 75 nmol mol(-1) ozone (O(3)) for 7 h d(-1) (O(3)-grown plants) for 28 d, and then exposed to 150 nmol mol(-1) O(3 )for 8 h. The concentration of ascorbate (ASC) was determined in leaf extracellular washing fluid (apoplast) and in the residual leaf tissue (symplast) after 0, 4 and 8 h acute fumigation, and after a 16 h "recovery" period in CFA. Changes in stomatal conductance were measured in vivo in order to model pollutant uptake, while the light-saturated rate of CO(2) assimilation (A:(sat)) was recorded as an indicator of O(3)-induced intracellular damage. Measurements of A:(sat) revealed enhanced tolerance to 150 nmol mol(-1) O(3) in plants pre-exposed to the pollutant compared with equivalent plants grown in CFA, consistent with the observed reduction in pollutant uptake due to lower stomatal conductance. The concentration of ASC in the leaf apoplast (ASC(apo)) declined upon O(3)-treatment in both CFA- and O(3)-grown plants, consistent with the oxidation of ASC(apo) under O(3)-stress. Furthermore, the decline in ASC(apo) was reversible in O(3)-grown plants after a 16 h "recovery" period, but not in plants grown in CFA. No significant change in the level and/or redox state of ASC in the symplast (ASC(symp)) was observed in plants exposed to 150 nmol mol(-1) O(3), and there was no difference in the constitutive level of ASC(symp) between CFA- and O(3)-grown plants. Model calculations indicated that the reaction of O(3) with ASC(apo) in the leaves of Vicia faba is potentially sufficient to intercept a substantial proportion (30-40%) of the O(3)entering the plant under environmentally-relevant conditions. The potential role of apoplastic ASC in mediating the tolerance of leaves to O(3) is discussed.

Air Pollutants↗

PubMed1987

Toxicity, radiation sensitivity modification, and combined drug effects of ascorbic acid with misonidazole in vivo on FSaII murine fibrosarcomas.

Ascorbic acid (ASC) has been shown to radioprotect nonmalignant tissue and to enhance the effects of misonidazole (MISO) on hypoxic cells in vitro. Since ASC is minimally toxic, it is an interesting candidate for improving the radiotherapeutic gain factor in vivo. The in vivo radiomodifying effects of ASC on a C3H/fSed murine fibrosarcoma (FSaII), on normal skin, and on bone marrow were determined with the use of the tumor growth delay and TCD50 (i.e., radiation dose required to control 50% of treated tumors for a minimum of 120 days) assays and the RD50 (i.e., dose required to cause a peak skin reaction of 2 in 50% of treated hind limbs) and LD50 (i.e., whole-body radiation lethal dose) assays, respectively. ASC was buffered to pH 7.35 and delivered ip at a dose of 4.5 mg g-1 body weight. ASC did not modify tumor growth delay induced by radiation or the TCD50 [87.1 Gy (control) vs. 85.6 Gy (ASC)]. Normal tissues, however, were radioprotected by ASC. RD50 values for 2+ acute skin reactions were 46.4 and 55.7 Gy for control and ASC-treated subjects, respectively; LD50 (30 days) values were 7.2 and 8.5 Gy. The enhancement ratios for skin and bone marrow were 1.20 and 1.18, and 95% confidence limits were (1.07 ... 1.34) and (1.14 ... 1.23), respectively. The therapeutic gain factor was 1.22 calculated as the ratio of the TCD50 and the reference normal tissue (RD50 or LD50). When ASC and MISO were combined, ASC reduced the in vivo radiosensitizing effects of MISO.

Animals↗

PubMed1995

Anti-CD3-activated splenocytes enhance survival in lethally irradiated mice after transplant of syngeneic hematopoietic stem cells.

Although cytokines produced by activated T cells may accelerate immunohematopoietic reconstitution after autologous bone marrow transplantation (ABMT), there is no direct evidence that infusion of anti-CD3 mAb-activated T cells can accelerate engraftment by hematopoietic stem cells. This study tests the ability of anti-CD3-activated murine splenocytes (ASC) to enhance the rescue of lethally irradiated (9 Gy) BDF1 mice by transplant of a limiting dose of fresh unmanipulated syngeneic splenocytes (SC). A minority (14.8%, 10-25%) of mice could be rescued with 5 x 10(5) SC after 9 Gy total-body irradiation (TBI). When 10(6) or 10(7) ASC were added to 5 x 10(5) SC, survival increased to 50% in those that received 5 x 10(5) SC + 10(6) ASC (not significant [NS]) and to 81.4% (77.7-88.0%) in those that received 5 x 10(5) SC + 10(7) ASC (p < 0.001). Furthermore, adding a fixed dose of 10(7) ASC to increasing doses of SC (10(5), 5 x 10(5), and 10(6)) enhanced survival at the different doses of SC. ASC alone did not rescue mice. CD3+ cells were the predominant population (77.6 +/- 6.7%) in the ASC inoculum, while NK cells remained low (1.2 +/- 0.9%). Colony-forming unit-spleen (CFU-S) yield after injection of SC showed dose dependence, whereas injection of 10 x 10(6) ASC alone failed to show any CFU-S yield in 23 of 25 recipient spleens. These results show that ASC enhanced survival of mice rescued with limiting doses of SC and that this effect was ASC dose-dependent but not dependent on the addition of extra stem cells.

Animals↗

PubMed1997

Homing potentials of circulating lymphocytes in humans depend on the site of activation: oral, but not parenteral, typhoid vaccination induces circulating antibody-secreting cells that all bear homing receptors directing them to the gut.

Specific Ab-secreting cells (ASC) appear in the human blood as a response to oral and parenteral vaccination. The actual contribution of these cells to the defense of the body depends on their final effector site. The homing potentials of mucosally and parenterally induced ASC were compared by examining the homing receptor (HR) expression of circulating specific ASC in the blood of volunteers vaccinated orally or parenterally with the same Ag, Salmonella typhi Ty21a. Circulating lymphocytes were separated into receptor-positive and -negative populations, and the numbers of specific ASC were assayed. The alpha4 beta7 integrin, which acts as a gut HR, was expressed on all (99%) of the mucosally activated ASC, but on only 58% of the parenterally induced ASC or 58% of all Ig-secreting cells of the unvaccinated controls. L-selectin, the peripheral lymph node HR, showed an inverse distribution; it was found on 42% of mucosally activated ASC and on 86% of parenterally induced ASC. These results reveal that all of the circulating ASC after oral vaccination are committed to migrate to the mucosal compartment of the immune system, strongly arguing for a recirculation of activated mucosal cells in humans. By contrast, ASC induced by parenteral vaccination with the same Ag are mostly directed to the systemic compartment, yet a part of them has mucosal homing attitudes as well. These differences indicate that the site of Ag encounter determines the homing potential of the cell.

Administration, Oral↗

PubMed2000

Activity of Se-allylselenocysteine in the presence of methionine gamma-lyase on cell growth, DNA integrity, apoptosis, and cell-cycle regulatory molecules.

Se-allylselenocysteine (ASC) is effective in inhibiting mammary epithelial cell growth in vitro and mammary carcinogenesis in vivo, but its mechanism is unknown. We recently reported that ASC reduces cell growth in a dose- and time-dependent manner, induces a loss of DNA integrity, and increases apoptosis. However, the level of ASC required for growth inhibition in vitro is 10- to 20-fold higher than that required in vivo. One possible explanation for this difference is that the cells used in in vitro studies have limited lyase activity required to release the allyl Se moiety from selenocysteine, whereas animals have abundant lyase activity in tissues. In the present study, we found that methionine gamma-lyase (MGL) added to culture medium containing ASC produced biological effects with lower levels of ASC, comparable to the selenium levels in plasma achieved during in vivo chemoprevention. The combination of 2.5 microM ASC and MGL inhibited the growth of TM12 cells and increased apoptosis without loss of DNA integrity. Treatment of TM12 cells with ASC and MGL resulted in an elevation of the protein levels of p53, Cip1/p21, and Kip1/p27, concomitant with a decrease in cyclins D1 and E and modest reductions in cyclin-dependent kinase inhibitors 4 and 2. Cells treated with ASC and MGL also showed decreased phosphorylation of retinoblastoma tumor-suppressor protein. Taken together, these results suggest that a physiologically relevant concentration of ASC with MGL exerts an inhibitory effect on cell growth and that this effect is likely to involve modulation of signaling pathways that suppress the phosphorylation of retinoblastoma tumor-suppressor protein.

Apoptosis↗

PubMed1990

Effect of phenytoin on the clinical pharmacokinetics of amiodarone.

Five healthy male volunteers were given oral amiodarone hydrochloride, 200 mg per day for 6 1/2 weeks, to determine its effects on the pharmacokinetics of both intravenous and oral phenytoin. Predose amiodarone and N-desethylamiodarone serum concentrations were obtained weekly during weeks 2-6. Amiodarone serum concentrations (ASC) increased during weeks 2-4 and then decreased sharply during weeks 5-6 when oral phenytoin, 2-4 mg/kg/day, was co-administered. In addition, N-desethylamiodarone serum concentrations (DEASC) exceeded corresponding ASC during weeks 5-6 whereas during weeks 2-4, DEASC were less than ASC. Because of the long elimination half-life for amiodarone previously reported in healthy volunteers after single doses of amiodarone and the frequent administration of amiodarone associated with this half-life, a modified equation for a continuous infusion was used to describe each subject's ASC versus time data. Pre-phenytoin ASC were fitted to an appropriate function to predict ASC during weeks 5-6 assuming no interaction. Observed versus predicted ASC were compared for weeks 5 and 6. Observed ASC during weeks 5 and 6 were (mean +/- SD) 0.25 +/- 0.09 micrograms/mL and 0.19 +/- 0.07 micrograms/mL, respectively. Corresponding predicted ASC were 0.36 +/- 0.12 micrograms/mL (P = .011) and 0.38 +/- 0.13 micrograms/mL (P = .004). These represented percent differences of 32.2 +/- 12.5% and 49.3 +/- 5.6% for weeks 5 and 6, respectively. Assuming there were no changes in the bioavailability of amiodarone during continuous administration, these findings strongly suggest induction of amiodarone metabolism by phenytoin. The clinical significance of this interaction remains to be determined.

Administration, Oral↗

PubMed1992

Inhibition of human term placental and fetal liver glutathione-S-transferases by fatty acids and fatty acid esters.

Glutathione-S-transferase (GST) activity from human term placenta and human fetal liver towards 1-chloro-2,4-dinitrobenzene as the second substrate was significantly inhibited by the saturated fatty acids, stearic (SA) and palmitic (PA) acids and fatty acid esters, ascorbyl stearate (Asc-S) and ascorbyl palmitate (Asc-P). The nature of inhibition of human placental GST was competitive towards CDNB with Ki values of 3.1, 10.0, 13.5 and 18.5 microM for Asc-S, Asc-P, PA and SA, respectively. The inhibitory effect of Asc-S on human term placental GST was reversible. I50 values for Asc-S, Asc-P, SA and PA were 15, 45, 83 and 78 microM, respectively, for partially purified human fetal liver GSTs and 21, 6, 88 and 117 microM, respectively, for partially pure rat liver GSTs. The evidence suggests that Asc-S, Asc-P, SA and PA are potent inhibitors especially of the pi-class of GST.

Animals↗

PubMed2005

Lack of the alanine-serine-cysteine transporter 1 causes tremors, seizures, and early postnatal death in mice.

The Na(+)-independent alanine-serine-cysteine transporter 1 (Asc-1) is exclusively expressed in neuronal structures throughout the central nervous system (CNS). Asc-1 transports small neutral amino acids with high affinity especially for D-serine and glycine (K(i): 8-12 microM), two endogenous glutamate co-agonists that activate N-methyl-D-aspartate (NMDA) receptors through interacting with the strychnine-insensitive glycine binding-site. By regulating D-serine (and possibly glycine) levels in the synaptic cleft, Asc-1 may play an important role in controlling neuronal excitability. We generated asc-1 gene knockout (asc-1(-/-)) mice to test this hypothesis. Behavioral phenotyping combined with electroencephalogram (EEG) recordings revealed that asc-1(-/-) mice developed tremors, ataxia, and seizures that resulted in early postnatal death. Both tremors and seizures were reduced by the NMDA receptor antagonist MK-801. Extracellular recordings from asc-1(-/-) brain slices indicated that the spontaneous seizure activity did not originate in the hippocampus, although, in this region, a relative increase in evoked synaptic responses was observed under nominal Mg(2+)-free conditions. Taken together with the known neurochemistry and neuronal distribution of the Asc-1 transporter, these results indicate that the mechanism underlying the behavioral hyperexcitability in mutant mice is likely due to overactivation of NMDA receptors, presumably resulting from elevated extracellular D-serine. Our study provides the first evidence to support the notion that Asc-1 transporter plays a critical role in regulating neuronal excitability, and indicate that the transporter is vital for normal CNS function and essential to postnatal survival of mice.

Amino Acid Transport System y+↗

PubMed2006

Biological principles for ex vivo adult stem cell expansion.

Adult stem cells (ASCs) are the engines that drive the renewal of adult mammalian tissues. They divide continuously, throughout life, to produce new progeny cells that undergo a robust development program of differentiation and maturation to replace older expired tissue cells. The same cell turnover program may function to provide limited repair and regeneration of adult tissues in some cases. The regenerative potential of ASCs drives the current intense interest in adapting them for applications in cell replacement therapy. However, research to explore this potential has been blunted by an unyielding biological problem. ASCs have proven highly refractory to expansion of their numbers and long-term propagation in culture. A review of reported strategies to overcome this problem reveals that many studies focus on traditional cell culture factors that may not apply to ASCs and overlook a special property of ASCs that may be universally critical for successful expansion, asymmetric cell kinetics (ACK). This property is reflected by the different kinetics fate of the two sister cells resulting from an ASC division: one cell remains an ASC and keeps the potential to divide for the entire life span of the tissue, while the other cell's progeny eventually differentiates and undergoes terminal division arrest. This unique property of ASCs may prove to be the obligatory factor that must be breached by any method that will succeed in accomplishing routine expansion of ASCs of diverse tissue origin.

Animals↗

PubMedDate not supplied

Circulating antibody secreting cell response to parenteral pneumococcal vaccines as an indicator of a salivary IgA antibody response.

This study assessed the mucosal immune response in healthy adult volunteers immunized parenterally with either pneumococcal polysaccharide (N = 8) or pneumococcal polysaccharide-protein conjugate (N = 10) vaccine with an aim to evaluate the relevance of antibody secreting cell (ASC) response after parenteral vaccination. An ASC response to the four types of capsular polysaccharide tested was observed in all vaccinees 7-9 days after immunization. IgA was the predominant class in the ASC response, and IgG the next common, with very few IgM ASCs. The IgA/IgG ratio in the ASC response was higher after immunization with the polysaccharide than the conjugate vaccine. Antibodies of the IgA class were frequently seen in the saliva already before immunization; especially to serotypes 14 and 19F. A twofold increase of the type specific secretory IgA antibodies in saliva was found in eight of the 16 instances in which the specific IgA ASC response was > 100 ASC per 10(6) cells and in only one of the 52 instances with fewer ASCs. We conclude that the ASC response in the peripheral blood is a useful parameter of the antibody response to pneumococcal vaccines and a good indicator of a secretory IgA response in the saliva.

Adult↗

PubMed2003

Atypical squamous cells: update on current concepts.

Besides all the confusion and associated problems that the use of ASC has created, it has initiated substantial investigational interest that has resulted in a better understanding of squamous intraepithelial lesions and the biology of cervical neoplasia. Although the category of ASC has created, and will continue to create, controversy in the diagnostic and management fields, it allows the pathologist to convey uncertainty that may be the result of poor sampling or difficulty in interpretation of a case. It is a valuable tool that the cytopathologist can use to make it known that the Papanicolaou test has its limitations and may need and benefit, in some instances, from support from ancillary studies. Similar limitations are recognized in other areas of pathology and the use of immunohistochemistry or molecular studies is widely accepted as an aid to a more specific and definitive interpretation. The time for the Papanicolaou test to be considered similarly has arrived. HPV DNA testing may not be the perfect test for cervical cancer screening because of high prevalence of HPV infection in the general population; however, it is currently the best-studied ancillary test and has been proven to be cost-effective for the triage of Papanicolaou tests with equivocal squamous cells. It is important for the cytopathologist to have well-developed diagnostic skills in interpreting gynecologic preparations, and to classify cases as ASC only when deemed appropriate. Downgrading cytologic findings that are diagnostic of a squamous intraepithelial lesion to ASC with the hope of supporting it by an HPV test will only result in a devaluation of the Papanicolaou test. Such recourse may, however, be acceptable in specific situations, such as in patients who have complex histories, atypical clinical presentations, or during pregnancy. Quality assurance measures to closely monitor the ASC:SIL ratio and the rate of HPV positivity in ASC cases will be essential to ensure the appropriate use of this interpretive category. The coordination of the 2001 Bethesda and ASCCP consensus meetings resulted in the new subcategories of ASC-US and ASC-H, along with well-defined management strategies for these interpretations. This new and clinically relevant terminology should lead to a reduction in difficulties at the clinical level and a more uniform management of patients, unlike the situation following Bethesda 1991 where the gynecologist was faced with a new "diagnosis" without specific management recommendations. The standardization of reporting and clinical management will also allow more reliable evaluation of patient outcomes and cost analysis. The 2002 American Cancer Society guidelines did not make specific recommendations regarding HPV DNA testing for the triage of patients who have a cytology result of ASC-US [64]. The FDA approved the expanded use of HPV testing in conjunction with the Papanicolaou test for cervical cancer screening in March of 2003. The future is likely to bring additional testing modalities that may be more specific for detecting squamous lesions that are more likely to persist or progress to carcinoma, than the currently available HPV tests. In addition, looking to the more distant future, recently published data from HPV vaccine trials suggests that immunizing women who are negative for HPV-16 may eventually reduce the incidence of cervical cancer [65]. At the present, however, the most effective method to decrease the mortality of this disease process is to make sure that all women have access to, and receive, effective cervical cytologic screening.

Female↗

PubMed2002

The effects of ascorbic acid on cartilage metabolism in guinea pig articular cartilage explants.

Ascorbic acid has been associated with the slowing of osteoarthritis progression in guinea pig and man. The goal of this study was to evaluate transcriptional and translational regulation of cartilage matrix components by ascorbic acid. Guinea pig articular cartilage explants were grown in the presence of L-ascorbic acid (L-Asc), D-isoascorbic acid (D-Asc), sodium L-ascorbate (Na L-Asc), sodium D-isoascorbate (Na D-Asc), or ascorbyl-2-phosphate (A2P) to isolate and analyze the acidic and nutrient effects of ascorbic acid. Transcription of type II collagen, prolyl 4-hydroxylase (alpha subunit), and aggrecan increased in response to the antiscorbutic forms of ascorbic acid (L-Asc, Na L-Asc, and A2P) and was stereospecific to the L-forms. Collagen and aggrecan synthesis also increased in response to the antiscorbutic forms but only in the absence of acidity. All ascorbic acid forms tended to increase oxidative damage over control. This was especially true for the non-nutrient D-forms and the high dose L-Asc. Finally, we investigated the ability of chondrocytes to express the newly described sodium-dependent vitamin C transporters (SVCTs). We identified transcripts for SVCT2 but not SVCT1 in guinea pig cartilage explants. This represents the first characterization of SVCTs in chondrocytes. This study confirms that ascorbic acid stimulates collagen synthesis and in addition modestly stimulates aggrecan synthesis. These effects are exerted at both transcriptional and post-transcriptional levels. The stereospecificity of these effects is consistent with chondrocyte expression of SVCT2, shown previously to transport L-Asc more efficiently than D-Asc. Therefore, this transporter may be the primary mechanism by which the L-forms of ascorbic acid enter the chondrocyte to control matrix gene activity.

Aggrecans↗

PubMed2002

Analysis of the entire nucleotide sequence of hepatitis B virus: characteristics of HBeAg-positive asymptomatic carriers, HBeAb positive asymptomatic carriers and patients with liver cirrhosis.

Entire nucleotide sequences of the HBV genome typical for various stages of HBV carriers are currently unknown. Comparison between conserved sequences in HBeAg-positive asymptomatic carriers (HBeAg ASCs) and mutations characteristic for HBeAb-positive asymptomatic carriers (HBeAb ASCs) are of clinical importance. In this study, we determined the entire nucleotide sequences of the HBV genome of patients infected with genotype C (HBeAg ASCs, 11 cases; HBeAb ASCs, seven cases; patients with liver cirrhosis (LC), five cases). Mutations in the entire nucleotide sequences were found more frequently in HBeAb ASCs than in HBeAg ASCs. In the precore/core (preC/C) region, amino acid mutations were more frequent in HBeAb ASCs (3.03%) than in HBeAg ASCs (0.00%) and patients with LC (0.69%). It was suggested that the mutations in the preC/C region had a close relationship with clinical status of HBV carriers. Mutations of leucine to isoleucine at a.a. 100 of the core region and of threonine to serine at a.a. 340 of the polymerase region were found frequently in HBeAb ASCs. In patients with LC, it was suggested that defective interfering particles (DI particles) play a role in the progression of stages. We conclude that attention should be given to mutations at a.a. 340 of the polymerase protein in addition to core protein.

Journal Article↗

PubMed2001

Ascorbate transport from the apoplast to the symplast in intact leaves.

Infiltration of reduced ascorbate (ASC) into the leaves of Betula pendula Roth and subsequent measurement of its loss therein after incubation allowed us to follow ascorbate transport from apoplast to symplast in intact leaves. All of the ascorbate extracted from the native apoplast was in fully oxidized form, dehydroascorbate (DHA). When 5 mM of ASC was infiltrated into the leaves, its intense decay occurred, but only 55% of ASC lost was recovered in apoplast as DHA. When ASC was added to the freshly extracted intercellular washing fluid (IWF), ASC oxidation occurred as well. However, all oxidized ASC was recovered as DHA, indicating that further decomposition of DHA did not occur. Similarly, all of the ASC infiltrated into the leaves was found therein either as ASC or DHA after incubation of leaves for up to 60 min. On this base the ascorbate infiltrated into the leaves and not recovered in the IWF was interpreted as ascorbate taken up into the symplast. The calculated uptake rates of ascorbate at different ASC concentrations followed saturation kinetics with the maximum uptake rate of 300 nmol m-2 plasma membrane (PM) area min-1 and Michaelis constant of 12.8 mM. The uptake of ascorbate was significantly inhibited by the addition of dithiothreitol or by PM H+ ATPase inhibitor erythrosin B. Thus, our results support the previous observations that DHA is preferably transported from the apoplastic to the cytoplasmic side of the membrane and show that this process is dependent upon PM proton gradient.

Journal Article↗

PubMed2000

A phase II study of two cycles of high-dose chemotherapy with autologous stem cell support in patients with metastatic breast cancer who meet eligibility criteria for a single cycle.

Multi-cycle high-dose chemotherapy with autologous stem cell support (HDC-ASCS) may improve the results obtained with single-cycle HDC-ASCS in metastatic breast cancer (MBC). However, the tolerability and efficacy of additional cycles of HDC-ASCS in patients selected using standard eligibility criteria for single cycle HDC-ASCS is uncertain. Twenty-nine patients with MBC and a CR or PR to induction chemotherapy were selected by standard institutional eligibility criteria for single-cycle HDC-ASCS. Cycle 1 HDC-ASCS (cyclophosphamide 6 g/m2; mitoxantrone 70 mg/m2; carboplatin 800 mg/m2) was followed by a planned second cycle (etoposide 1.6 g/m2; thiotepa 800 mg/m2; carboplatin 800 mg/m2 modulated by tamoxifen 120 mg/m2/day x 5 days) with a median interval of 3.2 months. CR rate was 20% after induction chemotherapy and 33% and 54% after HDC cycles I and II, respectively. Sixteen patients (55%) failed to complete HDC cycle II within 200 days because of disease progression, toxicity, inadequate stem cell collection, insurance denials or patient choice. Median progression-free survival (PFS) for all 29 patients entered is 301 days from date of HDC cycle I and actuarial PFS at 2 years is 35%. For the 13 patients who received the two cycles of HDC-ASCS, actuarial PFS at 2 years was 54% (P = NS compared to those receiving only one cycle). These data show that a second cycle of full-dose intensity HDC-ASCS may increase the proportion of patients with MBC that achieve CR and may increase PFS. However, a large proportion of patients that complete HDC-ASCS cycle I may fail to proceed to cycle II in a timely fashion. Bone Marrow Transplantation (2000) 25, 519-524.

Adult↗