PKA inhibition is a central step in D,L-methadone-induced ER Ca2+ release and subsequent apoptosis in acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) is a hematologic cancer that mostly affects children. It accounts for over a quarter of ALL pediatric cancers, causing most of the cancer death among children. Previously, we demonstrated that D,L-methadone causes ALL cell apoptosis via µ-opioid receptor 1 (OPRM1)-triggered ER Ca2+ release and decrease in Ca2+ efflux, elevating [Ca2+]i. However, the precise mechanism by which D,L-methadone induces ER Ca2+ release remains to be defined. Here, we show that in ALL cells, D,L-methadone-induced ER Ca2+ release is blocked by inhibition of Gαi, but not Gßϒ, indicating that the process is dependent on Gαi. Activation of adenylyl cyclase (AC) with forskolin or treatment with 8-CPT-cAMP blocks D,L-methadone-induced ER Ca2+ release, indicating that the latter results from Gαi-dependent downregulation of AC and cAMP. The 14-22 amide (myr) PKA inhibitor alone elicits ER Ca2+ release, and subsequent treatment with D,L-methadone does not cause additional ER Ca2+ release, indicating that PKA inhibition is a key step in D,L-methadone-induced ER Ca2+ release and can bypass the D,L-methadone-OPRM1-AC-cAMP step. This is consistent with the decrease in PKA-dependent (i) inhibitory PLCß3 Ser1105 phosphorylation that leads to PLCß3 activation and ER Ca2+ release, and (ii) BAD Ser118 phosphorylation, which together ultimately result in caspase activation and apoptosis. Thus, our findings indicate that D,L-methadone-induced ER Ca2+ release and subsequent apoptosis in ALL cells is mediated by Gαi-dependent downregulation of the AC-cAMP-PKA-PLCß3/BAD pathway. The fact that 14-22 amide (myr) alone effectively kills ALL cells suggests that PKA may be targeted for ALL therapy.

PKA inhibition kills L-asparaginase-resistant leukemic cells from relapsed acute lymphoblastic leukemia patients.

Despite the success in treating newly diagnosed pediatric acute lymphoblastic leukemia (aLL), the long-term cure rate for the 20% of children who relapse is poor, making relapsed aLL the primary cause of cancer death in children. By unbiased genome-wide retroviral RNAi screening and knockdown studies, we previously discovered opioid receptor mu 1 (OPRM1) as a new aLL cell resistance biomarker for the aLL chemotherapeutic drug, L-asparaginase, i.e., OPRM1 loss triggers L-asparaginase resistance. Indeed, aLL cell OPRM1 level is inversely proportional to L-asparaginase IC50: the lower the OPRM1 level, the higher the L-asparaginase IC50, indicating that aLL cells expressing reduced OPRM1 levels show resistance to L-asparaginase. In the current study, we utilized OPRM1-expressing and -knockdown aLL cells as well as relapsed patient aLL cells to identify candidate targeted therapy for L-asparaginase-resistant aLL. In OPRM1-expressing cells, L-asparaginase induces apoptosis via a cascade of events that include OPRM1-mediated decline in [cAMP]i, downregulation of PKA-mediated BAD S118 phosphorylation that can be reversed by 8-CPT-cAMP, cyt C release from the mitochondria, and subsequent caspase activation and PARP1 cleavage. The critical role of PKA inhibition due to a decrease in [cAMP]i in this apoptotic process is evident in the killing of OPRM1-knockdown and low OPRM1-expressing relapsed patient aLL cells by the PKA inhibitors, H89 and 14-22 amide. These findings demonstrate for the first time that PKA can be targeted to kill aLL cells resistant to L-asparaginase due to OPRM1 loss, and that H89 and 14-22 amide may be utilized to destroy L-asparaginase-resistant patient aLL cells.

Requirement for ER-mitochondria Ca2+ transfer, ROS production and mPTP formation in L-asparaginase-induced apoptosis of acute lymphoblastic leukemia cells.

Acute lymphoblastic leukemia (aLL) is a malignant cancer in the blood and bone marrow characterized by rapid expansion of lymphoblasts. It is a common pediatric cancer and the principal basis of cancer death in children. Previously, we reported that L-asparaginase, a key component of acute lymphoblastic leukemia chemotherapy, causes IP3R-mediated ER Ca2+ release, which contributes to a fatal rise in [Ca2+]cyt, eliciting aLL cell apoptosis via upregulation of the Ca2+-regulated caspase pathway (Blood, 133, 2222-2232). However, the cellular events leading to the rise in [Ca2+]cyt following L-asparaginase-induced ER Ca2+ release remain obscure. Here, we show that in acute lymphoblastic leukemia cells, L-asparaginase causes mitochondrial permeability transition pore (mPTP) formation that is dependent on IP3R-mediated ER Ca2+ release. This is substantiated by the lack of L-asparaginase-induced ER Ca2+ release and loss of mitochondrial permeability transition pore formation in cells depleted of HAP1, a key component of the functional IP3R/HAP1/Htt ER Ca2+ channel. L-asparaginase induces ER Ca2+ transfer into mitochondria, which evokes an increase in reactive oxygen species (ROS) level. L-asparaginase-induced rise in mitochondrial Ca2+ and reactive oxygen species production cause mitochondrial permeability transition pore formation that then leads to an increase in [Ca2+]cyt. Such rise in [Ca2+]cyt is inhibited by Ruthenium red (RuR), an inhibitor of the mitochondrial calcium uniporter (MCU) that is required for mitochondrial Ca2+ uptake, and cyclosporine A (CsA), an mitochondrial permeability transition pore inhibitor. Blocking ER-mitochondria Ca2+ transfer, mitochondrial ROS production, and/or mitochondrial permeability transition pore formation inhibit L-asparaginase-induced apoptosis. Taken together, these findings fill in the gaps in our understanding of the Ca2+-mediated mechanisms behind L-asparaginase-induced apoptosis in acute lymphoblastic leukemia cells.

Neuroprotective Effect of ß-secretase Inhibitory Peptide from Pacific Hake (Merluccius productus) Fish Protein Hydrolysate.

BACKGROUND: Various methodologies have been employed for the therapeutic interpolation of the progressive brain disorder Alzheimer's disease. Thus, ß-secretase inhibition is significant to prevent disease progression in the early stages. OBJECTIVE: This study seeks to purify and characterize a novel ß-secretase inhibitory peptide from Pacific hake enzymatic hydrolysate. METHODS: A potent ß-secretase inhibitory peptide was isolated by sequential purifications using Sephadex G-25 column chromatography and octadecylsilane (ODS) C18 reversed-phase HPLC. A total of seven peptides were synthesized using the isolated peptide sequences. SH-SY5Y cells stably transfected with the human ''Swedish'' amyloid precursor protein (APP) mutation APP695 (SH-SY5YAPP695swe) were used as an in-vitro model system to investigate the effect of Leu-Asn peptide on APP processing. RESULTS: The ß-secretase inhibitory activity (IC50) of the purified peptide (Ser-Leu-Ala-Phe-Val-Asp- Asp-Val-Leu-Asn) from fish protein hydrolysate was 18.65 µM and dipeptide Leu-Asn was the most potent ß-secretase inhibitor (IC50 value = 8.82 µM). When comparing all the seven peptides, the inhibition pattern of Leu-Asn dipeptide was found to be competitive by Lineweaver-Burk plot and Dixon plot (Ki value = 4.24 µM). The 24 h treatment with Leu-Asn peptide in SH-SY5Y cells resulted in reducing the ß-amyloid (Aß) production in a dose-dependent manner. CONCLUSION: Therefore, the results of this study suggest that ß-secretase inhibitory peptides derived from marine organisms could be potential candidates to develop nutraceuticals or pharmaceuticals as antidementia agents.

HAP1 loss confers l-asparaginase resistance in ALL by downregulating the calpain-1-Bid-caspase-3/12 pathway.

l-Asparaginase (l-ASNase) is a strategic component of treatment protocols for acute lymphoblastic leukemia (ALL). It causes asparagine deficit, resulting in protein synthesis inhibition and subsequent leukemic cell death and ALL remission. However, patients often relapse because of the development of resistance, but the underlying mechanism of ALL cell resistance to l-asparaginase remains unknown. Through unbiased genome-wide RNA interference screening, we identified huntingtin associated protein 1 (HAP1) as an ALL biomarker for l-asparaginase resistance. Knocking down HAP1 induces l-asparaginase resistance. HAP1 interacts with huntingtin and the intracellular Ca2+ channel, inositol 1,4,5-triphosphate receptor to form a ternary complex that mediates endoplasmic reticulum (ER) Ca2+ release upon stimulation with inositol 1,4,5-triphosphate3 Loss of HAP1 prevents the formation of the ternary complex and thus l-asparaginase-mediated ER Ca2+ release. HAP1 loss also inhibits external Ca2+ entry, blocking an excessive rise in [Ca2+]i, and reduces activation of the Ca2+-dependent calpain-1, Bid, and caspase-3 and caspase-12, leading to reduced number of apoptotic cells. These findings indicate that HAP1 loss prevents l-asparaginase-induced apoptosis through downregulation of the Ca2+-mediated calpain-1-Bid-caspase-3/12 apoptotic pathway. Treatment with BAPTA-AM [1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester)] reverses the l-asparaginase apoptotic effect in control cells, supporting a link between l-asparaginase-induced [Ca2+]i increase and apoptotic cell death. Consistent with these findings, ALL patient leukemic cells with lower HAP1 levels showed resistance to l-asparaginase, indicating the clinical relevance of HAP1 loss in the development of l-asparaginase resistance, and pointing to HAP1 as a functional l-asparaginase resistance biomarker that may be used for the design of effective treatment of l-asparaginase-resistant ALL.

Risk of Cardiovascular Disease Using Framingham Risk Score in Korean Cancer Survivors.

BACKGROUND: Cardiovascular disease is an important cause of morbidity and mortality in cancer survivors. The aim of this study was to investigate the modifiable cardiovascular disease risk factors and 10-year probability of the disease based on the Framingham risk score in cancer survivors, compared with the general population. METHODS: A total of 1,225 cancer survivors and 5,196 non-cancer controls who participated in the 2007-2013 Korea National Health and Nutrition Examination Surveys were enrolled. We assessed modifiable cardiovascular disease risk factors including smoking, body mass index, physical inactivity, high blood pressure, high cholesterol, and elevated blood glucose level. The 10-year probability of cardiovascular disease was determined by applying the Framingham cardiovascular disease risk equation among cancer survivors and non-cancer controls, ranging from 30 to 74 years old who had no overt cardiovascular diseases. RESULTS: The proportion of subjects who had higher fasting glucose levels, hemoglobin A1c levels, systolic blood pressure, and low density lipoprotein cholesterol levels, and those who had lower high density lipoprotein cholesterol levels was significantly higher in the cancer survivors than in the non-cancer controls. The average 10-year probability of cardiovascular disease among the cancer survivors was higher than that in the non-cancer controls in both men and women. The average 10-year probability of cardiovascular disease in relation to the cancer type was significantly higher in patients with hepatic, colon, lung, breast, and gastric cancer. CONCLUSION: Cancer survivors have a higher cardiovascular disease risk and 10-year probability of cardiovascular disease than non-cancer controls. Control of cardiovascular disease risk factors and implementation of a well-defined cardiovascular disease prevention program are needed for treating cancer survivors.

Association between Smoking and Periodontal Disease in Korean Adults: The Fifth Korea National Health and Nutrition Examination Survey (2010 and 2012).

BACKGROUND: This study aimed to evaluate an association between smoking, smoking cessation, and periodontal disease in Korean adults. METHODS: The data were collected from 8,336 participants, aged between 20 and 64 years, who participated in the fifth Korea National Health and Nutrition Examination (2010 and 2012). Smoking status was assessed using self-administered questionnaires. Periodontal disease was defined as a community periodontal index ≥3 points. Logistic regression analysis was used to evaluate an association between smoking, smoking cessation, and periodontal disease after adjusting for age, sex, education, monthly income, diabetes, obesity, alcohol intake, and frequency of tooth brushing. RESULTS: The risk of periodontal disease was higher among current smokers (odds ratio [OR], 1.49; 95% confidence interval [CI], 1.21-1.83) than never smokers. Among current smokers, the risk of periodontal disease was increased in smokers of ≥10 cigarettes/d, ≥20 years duration, and >10 pack-years compared with never smokers (P<0.05). Among former smokers, the risk of periodontal disease after 10 years since cessation declined to 0.56 (95% CI, 0.42-0.75) compared with current smokers and was indistinguishable statistically from never smokers. CONCLUSION: Periodontal disease is significantly associated with smoking status in Korean adults.

Factors Associated with Fatigue in Korean Gastric Cancer Survivors.

BACKGROUND: Gastric cancer is the second most common cancer in Korea. Fatigue is a common symptom among cancer survivors. The aim of this study was to identify factors associated with fatigue in gastric cancer survivors. METHODS: Data were analyzed from 199 gastric cancer survivors who visited a cancer survivor outpatient clinic from July 2013 to June 2014. Patients were surveyed using a questionnaire containing a fatigue severity scale (FSS) and questions regarding associated symptoms. Participants were divided into fatigue (FSS) and non-fatigue groups based on FSS scores (≥4 and <4, respectively). Age, sex, weight, body mass index, cancer stage, pathology, surgery type, chemotherapy, radiotherapy, comorbid disease, family history of cancer, smoking, alcohol consumption, exercise, and laboratory results were investigated. RESULTS: The fatigue and non-fatigue groups contained 42 and 157 survivors, respectively. Their mean age was 58 years, and the mean post-operative period was 6.58 years. Arthralgia (odds ratio [OR], 12.95; 95% confidence interval [CI], 3.21-52.34), dyspnea (OR, 10.54; 95% CI, 2.94-37.80), dyspepsia (OR, 8.26; 95% CI, 2.63-25.96), changed bowel habits (OR, 4.56; 95% CI, 1.09-19.11), anemia (OR, 3.18; 95% CI, 1.26-8.05), and regular exercise (OR, 0.31; 95% CI, 0.12-0.77) were significantly associated with fatigue in gastric cancer survivors, while weight, treatment, and depressive mood were not. CONCLUSION: Arthralgia, dyspnea, dyspepsia, bowel habit change, anemia, and regular exercise are associated with fatigue in gastric cancer survivors.

Health-Related Quality of Life and Cardiovascular Disease Risk in Korean Adults.

BACKGROUND: Health-related quality of life (HRQoL) is considered an important outcome measure in chronic diseases, in particular cardiovascular disease (CVD), which is known to be associated with impaired HRQoL. However, few studies have examined HRQoL in individuals at high risk of CVD. METHODS: Using the Fifth Korea National Health and Nutrition Examination Survey 2010-2012, we analyzed data from 10,307 adults aged ≥30 years. The study subjects were stratified into 3 groups on the basis of their Framingham risk score-a 10-year estimate of CVD risk: <10.0% (low risk), 10.0%-19.9% (moderate risk), and ≥20.0% (high risk). The EuroQol-5D (EQ-5D) was used to evaluate HRQoL. RESULTS: A significantly higher proportion of high-risk subjects than low-risk participants had impaired HRQoL (defined as the lowest quartile of the EQ-5D index); this held true even after adjustment for confounding factors in multivariable logistic regression analysis (men: odds ratio [OR], 1.62; 95% confidence interval [CI], 1.24-2.11; women: OR, 1.46; 95% CI, 1.02-2.08). In terms of the 5 EQ-5D dimensions, a 10-year CVD risk ≥20.0% was significantly associated with self-reported problems of mobility in men (OR, 3.15; 95% CI, 2.02-4.90), and of mobility (OR, 1.56; 95% CI, 1.09-2.24), self-care (OR, 2.14; 95% CI, 1.09-4.22), and usual activity problems (OR, 1.80; 95% CI, 1.17-2.78) in women. CONCLUSION: A high CVD risk is associated with impaired HRQoL. After adjustment for demographic and clinical factors, a 10-year CVD risk ≥20.0% is an independent predictor of impaired HRQoL in the general population; in particular, of mobility problems in men, and of mobility, self-care, and usual activity problems in women.

Chlorella ethanol extract induced phase II enzyme through NFE2L2 (nuclear factor [erythroid-derived] 2-like 2, NRF2) activation and protected ethanol-induced hepatoxicity.

In this study, we investigated the hepatoprotective effects of ethanol extracts from Chlorella vulgaris (CH) on animals. We measured its effect on the quinone reductase (QR) activity in Hepa1c1c7 cells, finding that CH induced a significantly higher QR activity in these cells. We isolated the active fraction (CH F4-2) from CH using chromatography methods. CH F4-2 may activate cellular antioxidant enzymes through upregulation of the Nrf2 pathway in hepatocarcinoma cells with CH F4-2 (25.0-200 µg/mL) for 48 h. Furthermore, CH F4-2 increased the expression of NQO1 [ NAD(P)H: quinone oxidoreductase, also known as QR], heme oxygenase-1, and glutathione-S-transferase P. Moreover, we found that ethanol-induced hepatic pathological changes-elevations in glutamic oxaloacetic transaminase, glutamic pyruvic transaminase, γ-glutamyltransferase, and lactate dehydrogenase-were significantly decreased. The inhibitory effect of CH on alcohol-induced liver injury was associated with the suppression of alcohol-induced increases in intestinal permeability. The ethanol extract from CH was found to induce QR activation, making it a potentially good candidate for a hepatoprotection agent.

Validation of the prognosis in palliative care study predictor models in terminal cancer patients.

BACKGROUND: Prognosis in Palliative Care Study (PiPS) predictor models were developed in 2011 to estimate the survival of terminal cancer patients in the United Kingdom. The aim of this study was to validate the PiPS model for terminal cancer patients in Korea, and evaluate its value in clinical practice. METHODS: This study included 202 advanced cancer patients who were admitted to the cancer hospital's palliative care ward from November 2011 to February 2013. On admission, physicians recorded the PiPS-A, PiPS-B, and doctor's survival estimates in inpatients. RESULTS: The median survival across PiPS-A categories was 9, 28, and 33 days, and the median survival across PiPS-B was 9.5, 27, and 43 days. The median actual survival was 25 days; overall accuracy between the PiPS-A, PiPS-B, doctor's estimates of survival, and actual survival was 52.0%, 49.5%, and 46.5%, respectively. The PiPS-A and PiPS-B groups for survival in 'days' showed a sensitivity of 48.4% and 64.1%, and specificity of 87.7%, and 77.5%, respectively. The PiPS-A and PiPS-B groups for survival in 'weeks' showed a sensitivity of 59.2%, and 44.7%, and specificity of 61.6%, and 64.7%, respectively. The PiPS-A and PiPS-B 'months' group showed a sensitivity of 37.1% and 37.1%, and specificity of 74.9% and 78.4%, respectively. The 'weeks' and 'months' groups showed significantly prolonged survival rates than 'days' group did in both PiPS-A and PiPS-B, by the Kaplan-Meier method. CONCLUSION: The PiPS predictor models effectively predicted the survival ≥14 days in terminal cancer patients, and were superior to doctor's estimates.

Association between Coffee Consumption and Renal Impairment in Korean Women with and without Diabetes: Analysis of the Fourth Korea National Health and Nutrition Examination Survey in 2008.

BACKGROUND: Recent studies suggest that coffee consumption has an influence on kidney function. This study investigated the relationship between habitual coffee consumption and renal impairment in Korean women, in consideration of diabetic status. METHODS: This study involved 2,673 women aged 35 to 84 years who had participated in the Fourth Korea National Health and Nutrition Examination Surveys, conducted in 2008. Habitual coffee consumption was classified into three categories: less than 1 cup per day, 1 cup per day, and 2 or more cups per day. Renal function impairment was defined as an estimated glomerular filtration rate less than 60 mL/min/1.73 m(2) by the Modification of Diet in Renal Disease equation. RESULTS: The prevalence of diabetes and renal function impairment was higher in women who drank < 1 cup of coffee per day. Compared with drinking < 1 cup of coffee per day, the odds ratio (OR) for renal function impairment was significantly lower (OR, 0.59; 95% confidence interval [CI], 0.37 to 0.95; P = 0.03) in those who habitually drank ≥ 2 cups per day after adjusting for multiple confounding factors. When data were stratified according to the presence of diabetes, coffee consumption ≥ 2 cups of coffee per day showed an inverse association with renal function impairment in only diabetic women (OR, 0.14; 95% CI, 0.02 to 0.88; P = 0.04), compared with consumption < 1 cup of coffee per day. CONCLUSION: In a representative sample of Korean women, coffee consumption was significantly associated with a decreased risk of renal impairment especially in middle and elderly-aged diabetic women.

ß-secretase inhibitory activity of phenolic acid conjugated chitooligosaccharides.

Eight kinds of phenolic acid conjugated chitooligosaccharides (COSs) were synthesized using hydroxyl benzoic acid and hydroxyl cinnamic acid. These phenolic acid conjugated-COSs with different substitution groups, including p-hydroxyl, 3,4-dihydroxyl, 3-methoxyl-4-hydroxyl and 3,5-dimethoxyl-4-hydroxy groups, were evaluated for their inhibitory activities against ß-site amyloid precursor protein (APP)-cleaving enzyme (BACE) and inhibited BACE with a ratio of 50.8%, 74.8%, 62.1%, 64.8% and 42.6%, respectively at the concentration of 1,000 µg/mL. BACE is a critical component to reduce the levels of Aß amyloid peptide in Alzheimer's disease (AD) which is based on the amyloid cascade theory in the brain, as this protease initiates the first step in Aß production. Among them, Caffeic acid conjugated-COS (CFA-COS) was further analysed to determine mode of inhibition of BACE and it showed non-competitive inhibition. Hence in this study, we suggest that CFA-COS derivatives have potential to be used as novel BACE inhibitors to reduce the risk of AD.

Nitric oxide-mediated vasorelaxation effects of anti-angiotensin I-converting enzyme (ACE) peptide from Styela clava flesh tissue and its anti-hypertensive effect in spontaneously hypertensive rats.

In our previous study, an anti-angiotensin I converting enzyme (ACE) peptide (Ala-His-Ile-Ile-Ile, MW: 565.3Da) was isolated from Styela clava flesh tissue. In this study the fractions obtained during the isolation process and the finally purified peptide were examined to see if they had vasorelaxation effects in isolated rat aortas, and then the peptide was investigated for anti-hypertensive effect in spontaneously hypertensive rats (SHRs). The induction of vasorelaxation in the rat aortas was observed with the isolated fractions and the peptide from the enzymatic hydrolysate of S. clava flesh tissue and could be markedly blocked by pretreatment with the nitric oxide synthase (NOS) inhibitor, N(G)-nitro-l-arginine methyl ester (l-NAME). In human endothelial cells, NO synthesis was found to be increased and eNOS phosphorylation was upregulated when the cells were cultured with the purified peptide. Furthermore, systolic blood pressure was reduced by administration of the potent vasorelaxation peptide in SHRs.

Characterization of bioactive peptides obtained from marine invertebrates.

Bioactive peptides as products of hydrolysis of diverse marine invertebrate (shellfish, crustacean, rotifer, etc.) proteins are the focus of current research. After much research on these muscles and by-products, some biologically active peptides were identified and applied to useful compounds for human utilization. This chapter reviews bioactive peptides from marine invertebrates in regarding to their bioactivities. Additionally, specific characteristics of antihypertensive, anti-Alzheimer, antioxidant, antimicrobial peptide enzymatic production, methods to evaluate bioactivity capacity, bioavailability, and safety concerns of peptides are reviewed.

Angiotensin I converting enzyme inhibitory peptide extracted from freshwater zooplankton.

In this study, hydrolysates obtained from the freshwater rotifer Brachionus calyciflonus were investigated for angiotensin I converting enzyme (ACE) inhibitory peptides. Freshwater rotifer protein was hydrolyzed using six separate enzymes in a batch reactor. The peptic hydrolysate had the highest ACE inhibitory activity compared to the other hydrolysates. The highest ACE inhibitory peptide was separated using Sephadex G-25 column chromatography and high-performance liquid chromatography on a C18 column. The 50% inhibitory concentration (IC(50)) value of purified ACE inhibitory peptide was 40.01 microg/mL. ACE inhibitory peptide was identified as being seven amino acid residues of Ala-Gln-Gly-Glu-Arg-His-Arg by N-terminal amino acid sequence analysis. The IC(50) value of purified ACE inhibitory peptide was 47.1 microM, and Lineweaver-Burk plots suggested that the peptide purified from rotifer protein acts as a competitive inhibitor against ACE. The results of this study suggest that peptides derived from freshwater rotifers may be beneficial as antihypertension compounds in functional foods or as pharmaceuticals.

Purification and characterization of angiotensin I converting enzyme inhibitory peptides from the rotifer, Brachionus rotundiformis.

Angiotensin I converting enzyme (ACE) inhibitory peptide was isolated from the marine rotifer, Brachionus rotundiformis. ACE inhibitory peptides were separated from rotifer hydrolysate prepared by Alcalase, alpha-chymotrypsin, Neutrase, papain, and trypsin. The Alcalase hydrolysate had the highest ACE inhibitory activity compared to the other hydrolysates. The IC(50) value of Alcalase hydrolysate for ACE inhibitory activity was 0.63 mg/ml. We attempted to isolate ACE inhibitory peptides from Alcalase prepared rotifer hydrolysate using gel filtration on a Sephadex G-25 column and high performance liquid chromatography on an ODS column. The IC(50) value of purified ACE inhibitory peptide was 9.64 microM, and Lineweaver-Burk plots suggest that the peptide purified from rotifer protein acts as a competitive inhibitor against ACE. Amino acid sequence of the peptide was identified as Asp-Asp-Thr-Gly-His-Asp-Phe-Glu-Asp-Thr-Gly-Glu-Ala-Met, with a molecular weight 1538 Da. The results of this study suggest that peptides derived from rotifers may be beneficial as anti-hypertension compounds in functional foods resource.

Results of a survey of residents who experience pregnancy and delivery.

PURPOSE: A growing number of residents bear children during residency training. This study was conducted to examine the pregnancy and maternity leave experiences of women who delivered infants during their residency training. METHODS: From March 2007 through June 2007, we surveyed 94 women who had given birth during their residency training during the past 5 years from the survey date. Participants were recruited using snowball sampling. RESULTS: Most of the respondents (70.2%) delivered in the 3rd and 4th year of residency. The average length of leave was 68.9 days, and more than half (54.3%) of the women expressed that a maternity leave policy was not formally established in the department. In general, participants believed that having a child during residency was difficult (mean=4.18 in 5-point Likert scale), and this perception was inversely related with systematic departmental support. CONCLUSION: Pregnancy is a special challenge for residents, with regard to not only physical overexertion but also psychosocial impacts on pregnant residents and their colleagues. However, most current residency programs do not have written policies on pregnancy. A formal policy and more flexible curriculum can help both the training program and the pregnant resident.

Dose-specific or dose-dependent effect of growth hormone treatment on the proliferation and differentiation of cultured neuronal cells.

OBJECTIVE: GH controls the proliferation of cartilage, fibroblasts or the differentiation of adipose and muscle tissue. However, the effect of GH on neuronal cells remains unknown. The present study was conducted to determine the proliferative or differentiating effect of GH on the nervous system in vitro. DESIGN: Neuronal hybrid cells (VSC4.1) were cultured with GH. The concentration ranged from 0.134 microg/ml up to 1.34 mg/ml. A cell confluency and MTT assay, cell cycle phase analysis with flow cytometry, extracellular receptor kinase (ERK) phosphorylation and mitogen activated protein kinase (MAPK) inhibitor (PD98050) assays were all performed to determine the effect on proliferation. Differentiation was evaluated by neurite outgrowth and neurofilament expression. Terminally differentiated neurons were stained by Hoechst 33342 for apoptotic nuclear fragmentation by degeneration. Poly-adenosyl ribose polymerase (PARP) expression and its cleavage products were evaluated. RESULTS: Cells at concentrations between 0.134 microg/ml and 1.34 microg/ml of GH proliferated with ERK phosphorylation, which was attenuated by MAPK inhibitors. Proliferation decreased at concentrations higher than 13.4 microg/ml; however, neurite outgrowth was observed at these concentrations. Terminally differentiated cells underwent apoptosis and showed nuclear fragmentation by Hoechst 33342 staining. PARP expression was increased with caspase-3 dependent-cleaved fragment. CONCLUSIONS: Our in vitro data demonstrate that GH exerts dual effects; proliferation with a specific GH dose window, or differentiation in a dose-dependent manner in cultured neuronal hybrid cells.