Measurement of Wall Shear Rate Across the Entire Vascular Wall Using Ultrasound Speckle Decorrelation.

OBJECTIVE: The accurate measurement of the wall shear rate (WSR) plays a crucial role in the early diagnosis of cardiovascular disease progression and acute events such as aneurysms and atherosclerotic plaque ruptures. To address this need, the speckle decorrelation (SDC) technique has been used to measure WSR based on the 2-D out-of-plane blood flow speed. This technique is particularly advantageous because it enables the use of a 1-D array transducer to measure WSR over the entire luminal area. This study aims to develop a region-based singular value decomposition (SVD) filtering technique that selectively suppresses clutter noise in the vascular region to measure WSR using SDC. METHOD: Ultrasound simulations, in-vitro flow experiments, and an in-vivo human study were conducted to evaluate the feasibility of this method's clinical application. RESULTS: The results demonstrated that WSR can be effectively measured across entire vascular walls using a conventional 1-D array transducer along with the proposed methodology. CONCLUSION: This study successfully demonstrates a noninvasive and accurate SDC-based method for measuring vital vascular WSR. This approach holds significant promise for assessing vascular WSR in both healthy individuals and high-risk cardiovascular disease patients.

Perioperative Management of Extended-Release Buprenorphine: A Narrative Review and Case Series.

BACKGROUND: Perioperative management of formulations of buprenorphine used for the treatment of opioid use disorder and/or pain are common clinical challenges. Care strategies are increasingly recommending continuation of buprenorphine while administering multimodal analgesia including full agonist opioids. While this "simultaneous strategy" is relatively simple for the shorter-acting sublingual buprenorphine formulation, best practices are needed for the increasingly prescribed extended-release buprenorphine (ER-buprenorphine). To our knowledge there are no prospective data to guide perioperative management of patients on ER-buprenorphine. Herein we provide a narrative review, report on the perioperative experiences of a series of patients maintained on ER-buprenorphine, and propose recommendations for perioperative ER-buprenorphine management based on best evidence, clinical experience, and our judgments. CASES: Here we present clinical data describing the perioperative experiences of patients maintained on extended-release buprenorphine who recently underwent a variety of surgeries ranging from outpatient inguinal hernia repair to multiple inpatient surgeries for source control in sepsis, at different medical centers throughout the United States. These patients were identified via an email solicitation to substance use disorder treatment providers throughout a nationwide healthcare system, requesting cases of patients maintained on extended-release buprenorphine who had recently undergone surgery. We report here on all of the cases received. DISCUSSION: Extrapolating from these and recently published case reports, we describe an approach to perioperative management of extended-release buprenorphine.

Ultrasound Speckle Decorrelation-Based Blood Flow Measurements.

Ultrasound imaging is the preferred noninvasive technique to measure blood flow to diagnose cardiovascular disease such as heart failure, carotid stenosis, and renal failure. Conventional ultrasound techniques such as Doppler ultrasound, ultrasound imaging velocimetry, vector Doppler and transverse oscillation beamforming have been used for blood flow velocity profile measurement. However, these techniques were limited to measuring blood flow velocities within the 2-D lateral (across the ultrasound beam) plane of a vessel, and the blood flow velocity profile was derived by assuming that blood vessels have a circular cross-section with axis symmetry. This assumption is incorrect because most vessels have complex geometries, such as tortuosity and branches, and an asymmetric flow profile in the presence of vascular plaque. Consequently, ultrasound speckle decorrelation has been proposed to measure blood flow from transverse views of blood vessels wherein the ultrasound beam is perpendicular to the vessel axis. In this review, we present a summary of recent progress in ultrasound speckle decorrelation-based blood flow measurement techniques.

Excellent activity and selectivity of Pd/ZSM-5 catalyst in the selective catalytic reduction of NOx by H2.

Superior de-NOx activity and N2 selectivity of the Pd/ZSM-5 catalyst was observed at low temperature (<200 °C) for the selective catalytic reduction of NOx by H2 (H2-SCR). Various Pd/ZSM-5 catalysts were prepared by calcinating at different temperatures (e.g., 500 °C, 650 °C, 750 °C, and 850 °C) and treated at reductive conditions before the H2-SCR reaction was performed. Among the prepared catalysts, the one prepared at the calcination temperature at 750 °C resulted in 96.7% NOx conversion and 96.8% N2 selectivity at 150 °C. Based on the H2-O2 reaction, the higher activity of the Pd/ZSM-5 catalyst calcined at 750 °C was attributed to its superior H2 activation ability for the H2-SCR reaction. The combined X-ray diffraction (XRD), temperature-programmed hydride decomposition (TPHD), and transmission electron microscopy (TEM) results revealed that highly dispersed Pd particles were generated on the catalyst calcined at 750 °C, while large Pd agglomerates were formed on the one calcined at 500 °C. It can be concluded that the catalytic activity of Pd/ZSM-5 improves by optimizing the calcination temperature, resulting in high Pd dispersion. Moreover, the Pd catalyst calcined at 750 °C showed high resistance to CO, maintaining >94% NOx conversion at 175 °C under 1000 ppm CO in the feed gas. Therefore, the catalyst calcined at 750 °C can be potentially used for industrial applications because of its simple preparation method and high resistance to CO.

Comparison of COVID-19 Pandemic-Related Stress among Frontline Medical Personnel in Daegu City, Korea.

Background and Objectives: Frontline medical staff usually experience high levels of stress, which could greatly impact their work output. We conducted a survey to investigate the level of stress and its association with job types, work departments, and medical centers among COVID-19 pandemic frontline medical personnel. Materials and Methods: We conducted a cross-sectional survey using a self-administered questionnaire among 307 frontline medical staff who cared for COVID-19 patients in Daegu city. We used a 33-item questionnaire to assess respondents' general characteristics, job stress, personal effects associated with the COVID-19 pandemic, and their stress level. A general health questionnaire-12 (GHQ-12) was included in our questionnaire. Results: Majority (74.3%) of the respondents were in the stress group. The mean GHQ-12 score was 14.31 ± 4.96. More females (67.4%, p < 0.05) and nurses (73.3%, p = 0.001) were in the stress group compared to males and doctors. Medical staff in the general ward considered the severity of the COVID-19 pandemic situation higher. Nurses perceived work changes (p < 0.05), work burden (p < 0.05), and personal impact (p < 0.05) more serious than doctors. Medical staff in Level 3 emergency department (ED) perceived a lack of real-time information (p = 0.012), a lack of resources, and negative personal impacts associated with the pandemic as more serious than staff in Level 1 and Level 2 EDs. Medical staff in the intensive care unit perceived work changes (p < 0.05), work burden (p < 0.05), and lack of personal protective equipment (p = 0.002) as more serious than staff in the ED and general ward. Conclusion: Providing real-time information and resources for reducing work burden and negative personal impact is central to maximizing the work output of the COVID-19 pandemic frontline medical staff. Supporting their mental health through regular programs and intervention is also imperative.

Laxative effects of triple fermented barley extracts (FBe) on loperamide (LP)-induced constipation in rats.

BACKGROUND: Constipation, a common health problem, causes discomfort and affects the quality of life. This study intended to evaluate the potential laxative effect of triple fermented barley (Hordeum vulgare L.) extract (FBe), produced by saccharification, Saccharomyces cerevisiae, and Weissella cibaria, on loperamide (LP)-induced constipation in Sprague-Dawley (SD) rats, a well-established animal model of spastic constipation. METHODS: Spastic constipation was induced via oral treatment with LP (3 mg/kg) for 6 days 1 h before the administration of each test compound. Similarly, FBe (100, 200 and 300 mg/kg) was orally administered to rats once a day for 6 days. The changes in number, weight, and water content of fecal, motility ratio, colonic mucosa histology, and fecal mucous contents were recorded. The laxative properties of FBe were compared with those of a cathartic stimulant, sodium picosulfate. A total of 48 (8 rats in 6 groups) healthy male rats were selected and following 10 days of acclimatization. Fecal pellets were collected one day before administration of the first dose and starting from immediately after the fourth administration for a duration of 24 h. Charcoal transfer was conducted after the sixth and final administration of the test compounds. RESULTS: In the present study, oral administration of 100-300 mg/kg of FBe exhibited promising laxative properties including intestinal charcoal transit ratio, thicknesses and mucous producing goblet cells of colonic mucosa with decreases of fecal pellet numbers and mean diameters remained in the lumen of colon, mediated by increases in gastrointestinal motility. CONCLUSION: Therefore, FBe might act as a promising laxative agent and functional food ingredient to cure spastic constipation, with less toxicity observed at a dose of 100 mg/kg.

Protective effects of triple fermented barley extract (FBe) on indomethacin-induced gastric mucosal damage in rats.

BACKGROUND: Hordeum vulgare L (barley) contains numerous phenolic substances with proven anticancer, antioxidant and gastroprotective activities. Saccharification increases the functionality and bioavailability of these compounds thus can aid in the development of a natural product based medicine. This study aimed to investigate the possible gastroprotective effects of saccharification on the indomethacin (IND)-induced gastric ulcers in rats using Weissella cibaria- and Saccharomyces cerevisiae-triple fermented H. vulgare extract (FBe). METHODS: In total, 60 healthy male 6-week old Sprague-Dawley SD (SPF/VAF Outbred CrljOri:CD1) rats were commercially purchased. The FBe extract (100, 200, and 300 mg kg- 1) was orally administered 30 min before an oral treatment of IND (25 mg kg- 1). Six hours after IND treatment, variations in the histopathology, myeloperoxidase (MPO) activity, gross lesion scores, lipid peroxidation, and antioxidant defense system component (superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH)) levels were measured. RESULTS: FBe treatment showed significant (p < 0.01 or p < 0.05) and dose-dependent decrease in gastric mucosal damage. In the present study hemorrhagic gross lesions, gastric MPO activity, and histopathological gastric ulcerative lesions were observed in IND-treated rats compared to the IND control rats. In particular, FBe, in a dose-dependent manner, strengthened the antioxidant defense systems, decreased lipid peroxidation and CAT activity by increasing the GSH levels and SOD activity, respectively. The 200 mg kg- 1 dose of FBe was similarly gastroprotective as the 10 mg kg- 1 dose of omeprazole in rats with IND-induced gastric mucosal damage. CONCLUSIONS: The findings of the present study show that an oral administration of FBe had positive gastroprotective effects through strengthening the body antioxidant defense system and anti-inflammatory effects.

Protective effects of a triple-fermented barley extract (FBe) against HCl/EtOH-induced gastric mucosa damage in mice.

This study was designed to observe the possible protective effects of a triple-fermented barley (Hordeum vulgare L.) extract (FBe) obtained by saccharification and using Saccharomyces cerevisiae and Weissella cibaria in alleviating gastric damage induced by a hydrochloric acid (HCl) and ethanol (EtOH) mixture in mice. After oral administration of FBe (300, 200, and 100 mg/kg) followed by 1 hr before and after the single treatment of HCl/EtOH (H/E) mixture, the hemorrhagic lesion scores, histopathology of the stomach, gastric nitrate/nitrite content, lipid peroxidation, and antioxidant defense systems including catalase and superoxide dismutase activities were observed. Following a single oral treatment of H/E-induced gastric damages as measured by hemorrhagic gross lesions and histopathological gastric, ulcerative lesions were significantly and dose-dependently (p < 0.01 or p < 0.05) inhibited in mice, when all three different doses of FBe were administered as compared to those in H/E control mice. In particular, FBe also increased gastric nitrate/nitrite content and strengthened the antioxidant defense, with a decrease in the level of gastric lipid peroxidation, but increased the activities of CAT and SOD. Moreover, the effects of FBe are comparable to that of ranitidine, a reference drug. The obtained results suggest that this fermented barley extract prevented mice from H/E-induced gastric mucosal damages through the suppression of inflammatory responses and oxidative stress-responsive free radicals. Thus, FBe can be useful to treat patients suffering from gastric mucosal disorders as a potent food supplement, and thereby, it would increase the necessity of application in the food industry.

Anti-osteoporotic effects of mixed compositions of extracellular polymers isolated from Aureobasidium pullulans and Textoria morbifera in ovariectomized mice.

BACKGROUND: Extracellular polymeric substances isolated from Aureobasidium pullulans (EAP), containing specifically 13% ß-1,3/1,6-glucan, have shown various favorable bone-preserving effects. Textoria morbifera Nakai (TM) tree has been used as an ingredient in traditional medicine and tea for various pharmacological purposes. Thus, the present study was aimed to examine the synergistic anti-osteoporotic potential of mixtures containing different proportions of EAP and TM compared with that of the single formulations of each herbal extract using bilateral ovariectomized (OVX) mice, a renowned rodent model for studying human osteoporosis. METHODS: Thirty five days after bilateral-OVX surgery, 9 combinations of EAP:TM (ratios = 1:1, 1:3, 1:5, 1:7, 1:9, 3:1, 5:1, 7:1, 9:1) and single separate formulations of EAP or TM were supplied orally, once a day for 35 days at a final concentration of 200 mg/kg. Variations in body weight gains during the experimental periods, as well as femur weights, bone mineral density (BMD), bone strength (failure load), and mineral content (calcium [Ca] and inorganic phosphorus [IP]) following sacrifice were measured. Furthermore, histomorphometric and histological profile analyses of serum biochemical parameters (osteocalcin content and bone specific alkaline phosphatase [bALP] activity) were conducted following sacrifice. Femurs histomorphometric analyses were also conducted for bone resorption, structure and mass. The results for the mixed formulations of EAP:TM and separate formulations were compared with those of risedronate sodium (RES). RESULTS: The EAP:TM (3:1) formulation synergistically enhanced the anti-osteoporotic potential of individual EAP or TM formulations, possibly due to enhanced variety of the active ingredients. Furthermore, the effects of EAP:TM were comparable to those of RES (2.5 mg/kg) treatment. CONCLUSION: The results of this study suggest that, the EAP:TM (3:1) combination might act as a new pharmaceutical agent and/or health functional food substance for curing osteoporosis in menopausal women.

Ingestion of Exopolymers from Aureobasidium pullulans Reduces the Duration of Cold and Flu Symptoms: A Randomized, Placebo-Controlled Intervention Study.

AIM: The objective of the study was to assess the efficacy of exopolymers from Aureobasidium pullulans (EAP) on the incidence of colds and flu in healthy adults. METHODS: We conducted a randomized, double-blind, placebo-controlled study at the onset of the influenza season. A total of 76 subjects (30-70 years of age) were recruited from the general population. The subjects were instructed to take one capsule per day of either EAP or a placebo for a period of 8 weeks. The duration of cold and flu symptoms, a primary variable in assessing effectiveness, and serum cytokine levels as well as WBC counts as secondary variables were also evaluated. RESULTS: EAP was associated with a statistically significant decrease in the duration of cold and flu symptoms, a primary variable in assessing effectiveness. Although cold and flu symptom levels were not significantly different at a significance level of 5%, the cold and flu symptom levels of the EAP group were less severe compared to the placebo group. No statistically significant changes of serum cytokine levels as well as WBC counts were observed. CONCLUSION: The results showed that EAP is a useful pharmaceutical and functional food material for preventing and treating colds and flu.

Extracellular polysaccharides purified from Aureobasidium pullulans SM­2001 (Polycan) inhibit dexamethasone­induced muscle atrophy in mice.

The present study assessed the beneficial skeletal muscle­preserving effects of extracellular polysaccharides from Aureobasidium pullulans SM­2001 (Polycan) (EAP) on dexamethasone (DEXA)­induced catabolic muscle atrophy in mice. To investigate whether EAP prevented catabolic DEXA­induced muscle atrophy, and to examine its mechanisms of action, EAP (100, 200 and 400 mg/kg) was administered orally, once a day for 24 days. EAP treatment was initiated 2 weeks prior to DEXA treatment (1 mg/kg, once a day for 10 days) in mice. Body weight alterations, serum biochemistry, calf thickness, calf muscle strength, gastrocnemius muscle thickness and weight, gastrocnemius muscle antioxidant defense parameters, gastrocnemius muscle mRNA expression, histology and histomorphometry were subsequently assessed. After 24 days, DEXA control mice exhibited muscle atrophy according to all criteria indices. However, these muscle atrophy symptoms were significantly inhibited by oral treatment with all three doses of EAP. Regarding possible mechanisms of action, EAP exhibited favorable ameliorating effects on DEXA­induced catabolic muscle atrophy via antioxidant and anti­inflammatory effects; these effects were mediated by modulation of the expression of genes involved in muscle protein synthesis (AKT serine/threonine kinase 1, phosphatidylinositol 3­kinase, adenosine A1 receptor and transient receptor potential cation channel subfamily V member 4) and degradation (atrogin­1, muscle RING­finger protein­1, myostatin and sirtuin 1). Therefore, these results indicated that EAP may be helpful in improving muscle atrophies of various etiologies. EAP at 400 mg/kg exhibited favorable muscle protective effects against DEXA­induced catabolic muscle atrophy, comparable with the effects of oxymetholone (50 mg/kg), which has been used to treat various muscle disorders.

Low molecular weight fucoidan improves endoplasmic reticulum stress-reduced insulin sensitivity through AMP-activated protein kinase activation in L6 myotubes and restores lipid homeostasis in a mouse model of type 2 diabetes.

Low molecular weight fucoidan (LMWF) is widely used to treat metabolic disorders, but its physiologic effects have not been well determined. In the present study, we investigated the metabolic effects of LMWF in obese diabetic mice (leptin receptor-deficient db/db mice) and the underlying molecular mechanisms involved in endoplasmic reticulum (ER) stress-responsive L6 myotubes. The effect of LMWF-mediated AMP-activated protein kinase (AMPK) activation on insulin resistance via regulation of the ER stress-dependent pathway was examined in vitro and in vivo. In db/db mice, LMWF markedly reduced serum glucose, triglyceride, cholesterol, and low-density lipoprotein levels, and gradually reduced body weights by reducing lipid parameters. Furthermore, it effectively ameliorated glucose homeostasis by elevating glucose tolerance. In addition, the phosphorylation levels of AMPK and Akt were markedly reduced by ER stressor, and subsequently, glucose uptake and fatty acid oxidation were also reduced. However, these adverse effects of ER stress were significantly ameliorated by LMWF. Finally, in L6 myotubes, LMWF markedly reduced the ER stress-induced upregulation of the mammalian target of rapamycin-p70S61 kinase network and subsequently improved the action of insulin via AMPK stimulation. Our findings suggest that AMPK activation by LMWF could prevent metabolic diseases by controlling the ER stress-dependent pathway and that this beneficial effect of LMWF provides a potential therapeutic strategy for ameliorating ER stress-mediated metabolic dysfunctions.

Glyceollin improves endoplasmic reticulum stress-induced insulin resistance through CaMKK-AMPK pathway in L6 myotubes.

Glyceollin has been shown to have antidiabetic properties, although its molecular mechanism is not known. Here, we have investigated the metabolic effects of glyceollin in animal models of insulin resistance and in endoplasmic reticulum (ER) stress-responsive muscle cells. db/db mice were treated with glyceollin for 4weeks and triglycerides, total cholesterol, low-density lipoprotein (LDL) and high-density lipoprotein (HDL) levels were measured. Glyceollin reduced serum insulin and triglycerides and increased HDL levels in db/db mice. Furthermore, glyceollin caused a significant improvement in glucose homeostasis without altering body weight and food intake in db/db mice. In muscle cells, glyceollin increased the activity of AMP-activated protein kinase (AMPK) as well as cellular glucose uptake. Fatty acid oxidation was also increased. In parallel, phosphorylation of acetyl-CoA carboxylase (ACC) at Ser-79 was increased, consistent with decreased ACC activity. An insulin-resistant state was induced by exposing cells to 5µg/ml of tunicamycin as indicated by decreased insulin-mediated tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) and glucose uptake. Inhibition of insulin-mediated tyrosine phosphorylation of IRS-1 and glucose uptake under ER stress was prevented by glyceollin. Strikingly, glyceollin reduced ER stress-induced, c-Jun NH2-terminal kinase activation and subsequently increased insulin signaling via stimulation of AMPK activity in L6 myotubes. Pharmacologic inhibition or knockdown of Ca(2+)/calmodulin-dependent protein kinase kinase blocked glyceollin-increased AMPK phosphorylation and insulin sensitivity under ER stress conditions. Taken together, these results indicate that glyceollin-mediated enhancement of insulin sensitivity under ER stress conditions is predominantly accomplished by activating AMPK, thereby having beneficial effects on hyperglycemia and insulin resistance.

Heat-processed Gynostemma pentaphyllum extract improves obesity in ob/ob mice by activating AMP-activated protein kinase.

Gynostemma pentaphyllum is widely used in Asian countries as a herbal medicine to treat dyslipidemia, type 2 diabetes and inflammation. An ethanol extract of G. pentaphyllum lessened obesity by activating AMP-activated protein kinase (AMPK). The levels of damulins A and B, components responsible for AMPK activation in the extract, were increased by autoclaving in a time-dependent manner. Heat-processed G. pentaphyllum extract, actiponin containing damulins A (0.93 %, w/w) and B (0.68 %, w/w), significantly stimulated fat oxidation and glucose uptake via AMPK activation in L6 myotube cells. Oral administration of actiponin to ob/ob mice for 8 weeks decreased body weight gain, liver weight, and blood cholesterol levels with AMPK activation in the soleus muscle. Our results demonstrate the beneficial effect of G. pentaphyllum on improving obesity and have elucidated the underlying molecular mechanisms.

New dammarane-type glucosides as potential activators of AMP-activated protein kinase (AMPK) from Gynostemma pentaphyllum.

AMP-activated protein kinase (AMPK) is a key sensor and regulator of glucose, lipid, and energy metabolism throughout the body. Activation of AMPK improves metabolic abnormalities associated with metabolic diseases including obesity and type-2 diabetes. The oriental traditional medicinal herbal plant, Gynostemma pentaphyllum, has shown a wide range of beneficial effects on glucose and lipid metabolism. In this study, we found that G. pentaphyllum contains two novel dammarane-type saponins designated as damulin A (1), 2α,3ß,12ß-trihydroxydammar-20(22)-E,24-diene-3-O-[ß-D-glucopyranosyl-(1→2)-ß-D-glucopyranoside], and damulin B (2), 2α,3ß,12ß-trihydroxydammar-20,24-diene-3-O-[ß-D-glucopyranosyl-(1→2)-ß-D-glucopyranoside], that strongly activate AMPK in cultured L6 myotube cells. Damulins A and B also increased ß-oxidation and glucose uptake with increasing GluT4 translocation to the plasma membrane in L6 myotube cells. Taken together our results indicate that activation of AMPK by damulins A and B may contribute to beneficial effect of G. pentaphyllum on glucose and lipid metabolism.

Antitumor Effects and Immunomodulating Activities of Phellinus linteus Extract in a CT-26 Cell-Injected Colon Cancer Mouse Model.

The antitumor effects of Phellinus linteus extract (Keumsa Linteusan) were investigated in a CT-26 cell-injected colon cancer mouse model. When administered orally (250~1,000 mg/kg body weight), Keumsa Linteusan significantly inhibited the growth of solid colon cancer. The highest dose was highly effective, reducing tumor formation by 26% compared with the control group. The anticomplementary activity of Keumsa Linteusan increased in a dose-dependent manner. Lysosomal enzyme activity of macrophages was increased by 2-fold (100 µg/ml) compared with the control group. Keumsa Linteusan can be regarded as a potent enhancer of the innate immune response, and can be considered as a very promising candidate for antitumor action.

Oral Single Dose Toxicity Study of Low Molecular Fucoidan in Mice.

This study was conducted to obtain information of the oral dose toxicity of low molecular fucoidan (LMF) in male and female mice. In order to calculate 50% lethal dose (LD50) and approximate lethal dose (LD), test material was once orally administered to male and female ICR mice at dose levels of 2000, 1000, 500, 250, 125 and 0 (vehicle control) mg/kg (body wt.). The mortality and the changes on body weight, clinical signs, gross observation and organ weight and histopathology of principle organs were monitored 14 days after LMF treatment. We could not find any mortalities, clinical signs, body weight changes and gross findings. In addition, significant changes in the organ weight and histopathology of principal organs were not observed except for some sporadic findings. The results obtained in this study suggest that LMF may not be toxic in mice and may be therefore safe for clinical use. The LD50 and approximate LD in mice after single oral dose of LMF were considered over 2000 mg/kg in both female and male mice.

Cytosolic NADP+-dependent isocitrate dehydrogenase plays a key role in lipid metabolism.

NADPH is an essential cofactor for many enzymatic reactions including glutathione metabolism and fat and cholesterol biosynthesis. We have reported recently an important role for mitochondrial NADP(+)-dependent isocitrate dehydrogenase in cellular defense against oxidative damage by providing NADPH needed for the regeneration of reduced glutathione. However, the role of cytosolic NADP(+)-dependent isocitrate dehydrogenase (IDPc) is still unclear. We report here for the first time that IDPc plays a critical role in fat and cholesterol biosynthesis. During differentiation of 3T3-L1 adipocytes, both IDPc enzyme activity and its protein content were increased in parallel in a time-dependent manner. Increased expression of IDPc by stable transfection of IDPc cDNA positively correlated with adipogenesis of 3T3-L1 cells, whereas decreased IDPc expression by an antisense IDPc vector retarded adipogenesis. Furthermore, transgenic mice with overexpressed IDPc exhibited fatty liver, hyperlipidemia, and obesity. In the epididymal fat pads of the transgenic mice, the expressions of adipocyte-specific genes including peroxisome proliferator-activated receptor gamma were markedly elevated. The hepatic and epididymal fat pad contents of acetyl-CoA and malonyl-CoA in the transgenic mice were significantly lower, whereas the total triglyceride and cholesterol contents were markedly higher in the liver and serum of transgenic mice compared with those measured in wild type mice, suggesting that the consumption rate of those lipogenic precursors needed for fat biosynthesis must be increased by elevated IDPc activity. Taken together, our findings strongly indicate that IDPc would be a major NADPH producer required for fat and cholesterol synthesis.

Cytosolic NADP(+)-dependent isocitrate dehydrogenase status modulates oxidative damage to cells.

NADPH is an important cofactor in many biosynthesis pathways and the regeneration of reduced glutathione, critically important in cellular defense against oxidative damage. It is mainly produced by glucose 6-phosphate dehydrogenase (G6PD), malic enzyme, and the cytosolic form of NADP(+)-dependent isocitrate dehydrogenase (IDPc). Little information is available about the role of IDPc in antioxidant defense. In this study we investigated the role of IDPc against cytotoxicity induced by oxidative stress by comparing the relative degree of cellular responses in three different NIH3T3 cells with stable transfection with the cDNA for mouse IDPc in sense and antisense orientations, where IDPc activities were 3-4-fold higher and 35% lower, respectively, than that in the parental cells carrying the vector alone. Although the activities of other antioxidant enzymes, such as superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase, and G6PD, were comparable in all transformed cells, the ratio of GSSG to total glutathione was significantly higher in the cells expressing the lower level of IDPc. This finding indicates that IDPc is essential for the efficient glutathione recycling. Upon transient exposure to increasing concentrations of H(2)O(2) or menadione, an intracellular source of free radicals and reactive oxygen species, the cells with low levels of IDPc became more sensitive to oxidative damage by H(2)O(2) or menadione. Lipid peroxidation, oxidative DNA damage, and intracellular peroxide generation were higher in the cell-line expressing the lower level of IDPc. However, the cells with the highly over-expressed IDPc exhibited enhanced resistance against oxidative stress, compared to the control cells. This study provides direct evidence correlating the activities of IDPc and the maintenance of the cellular redox state, suggesting that IDPc plays an important role in cellular defense against oxidative stress.