Medical biochemistry and metabolomics

The use of flow cytometry in the clinical laboratory has grown substantially in the past decade. This is attributable in part to the development of smaller, user-friendly, less-expensive instruments and a continuous increase in the number of clinical applications. Flow cytometry measures multiple characteristics of individual particles flowing in single file in a stream of fluid. Light scattering at different angles can distinguish differences in size and internal complexity, whereas light emitted from fluorescently labeled antibodies can identify a wide array of cell surface and cytoplasmic antigens. This approach makes flow cytometry a powerful tool for detailed analysis of complex populations in a short period of time. This report reviews the general principles in flow cytometry and selected applications of flow cytometry in the clinical hematology laboratory.

Background: Dietary diversity is a proxy indicator of nutrient adequacy. However, little is documented on dietary diversity among pregnant women in Ethiopia in general and specifically in the study area. This study assessed dietary diversity and associated factors among pregnant women attending antenatal care in public health facilities in Bale Zone, Southeast Ethiopia. Methods: An institution-based cross-sectional study was conducted in Bale Zone from January to March 2017. The sample size was determined using a single population proportion formula. Data were collected by pretested structured interviewer-administered questionnaires from a total of 413 pregnant women who were identified through systematic random sampling. The sample was drawn proportionally from selected public health facilities based on the client load. Dietary diversity was computed from information about the nine food groups obtained using a 24-hour dietary recall method. Statistical analysis was done using bivariate and multivariate logistic regression with the P-value <0.05 at 95% confidence interval considered as statistically significant. Results: The mean age of the pregnant women was 26.93 with standard deviation ±6.12 years. About 55.2% of the pregnant women had inadequate dietary diversity. Getting information from a health professional [AOR =5.26, 95% CI (1.60, 17.36)], being an urban dweller [AOR =8.95, 95% CI (4.42, 18.16)], having a protected water source [AOR =11.16, 95% CI (4.74, 26.27)], having a latrine [AOR =8.21, 95% CI (4.01, 16.80)], having a home garden [AOR =4.26, 95% CI (2.08, 8.70)], having a bank account [AOR =12.25, 95% CI (6.01, 24.97)] and having use of a mobile phone [AOR =3.82, 95% CI (1.92, 7.62)] were significantly associated with dietary diversity. Conclusion: In this community, the prevalence of inadequate dietary diversity is high. Variables which indicate a better living condition such as having a protected source of water, having a latrine, having a home garden, being an urban dweller, having a bank account and having use of a mobile phone were independent predictors of dietary diversity. Therefore, attention should be paid to improve to better living conditions of pregnant women by addressing determinate variables through community awareness.

The liking and selective ingestion of palatable foods-including sweets-is biologically controlled, and dysfunction of this regulation may promote unhealthy eating, obesity, and disease. The hepatokine fibroblast growth factor 21 (FGF21) reduces sweet consumption in rodents and primates, whereas knockout of Fgf21 increases sugar consumption in mice. To investigate the relevance of these findings in humans, we genotyped variants in the FGF21 locus in participants from the Danish Inter99 cohort (n = 6,514) and examined their relationship with a detailed range of food and ingestive behaviors. This revealed statistically significant associations between FGF21 rs838133 and increased consumption of candy, as well as nominal associations with increased alcohol intake and daily smoking. Moreover, in a separate clinical study, plasma FGF21 levels increased acutely after oral sucrose ingestion and were elevated in fasted sweet-disliking individuals. These data suggest the liver may secrete hor...

Obesity in humans is a major public health crisis worldwide. In addition, livestock species exhibit excessive subcutaneous fat at market weight. However, there are currently few means of reducing adiposity in mammals. This study was conducted with a swine model to test the hypothesis that dietary L-arginine supplementation may increase muscle gain and decrease fat deposition. Twenty-four 110-day-old barrows were assigned randomly into two treatments, representing supplementation with 1.0% L-arginine or 2.05% L-alanine (isonitrogenous control) to a corn-and soybean meal-based diet. Growth performance was measured based on weight gain and food intake. After a 60-day period of supplementation, carcass and muscle composition were measured. Serum triglyceride concentration was 20% lower (P \ 0.01) but glucagon level was 36% greater (P \ 0.05) in arginine-supplemented than in control pigs. Compared with the control, arginine supplementation increased (P \ 0.05) body weight gain by 6.5% and carcass skeletal-muscle content by 5.5%, while decreasing (P \ 0.01) carcass fat content by 11%. The arginine treatment enhanced (P \ 0.05) longissimus dorsi muscle protein, glycogen, and fat contents by 4.8, 42, and 70%, respectively, as well as muscle pH at 45 min post-mortem by 0.32, while reducing muscle lactate content by 37%. These results support our hypothesis that dietary arginine supplementation beneficially promotes muscle gain and reduces body fat accretion in growingfinishing pigs. The findings have a positive impact on development of novel therapeutics to treat human obesity and enhance swine lean-tissue growth.

To elucidate the role of endogenous hormones like testosterone, progesterone, estrogen, insulinlike growth factor, insulin and glucocorticoids in a common skin condition acne vulgaris. Design and methods: We conducted a systematic review of the literature and abstracted the data for every published cut point. Results: We screened more than 1000 studies and found that serum testosterone, progesterone, glucocorticoids, insulin and insulin-like growth factors are increased in patients with acne vulgaris and serum estrogen levels are low in patients. Conclusions: Various endogenous hormones play important role in the pathogenesis of acne vulgaris. Thus, in the clinical practice it is important to evaluate serum levels of these hormones and patients must be treated accordingly to avoid serious endocrine disorders at an early age.

Controversy still rages about whether hormone replacement therapy (HRT) confers cardiovascular benefit or harm. There is a wealth of biological evidence that estrogen has a beneficial effect, supporting a large body of epidemiological evidence demonstrating reduction in coronary events with HRT. A large randomized placebocontrolled clinical trial of preventive strategies for coronary heart disease (CHD) in postmenopausal women, the Women's Health Initiative (WHI), included HRT arms. The published preliminary findings of this trial showed a significant increase in coronary events, stroke, venous thromboembolism and breast cancer with estrogen-progestogen, leading to the conclusion that HRT was unsafe to use other than for short-term relief of menopausal symptoms. But subsequent publications of the more complete data from WHI have shown no significant increase in CHD, and a tendency to a reduction in those initiating HRT below age 60 years. This is important because other therapeutic strategies for the primary prevention of CHD, such as aspirin and statins, are not of proven benefit in women, in contrast to men. Subsequent WHI findings have not shown a clear increase in breast cancer, and any potential increase from HRT is similar to that seen with many lifestyle factors and other commonly used medications. The preliminary WHI results do not reflect accurately true benefits and risks, and HRT should remain a potential preventive treatment for CHD. J C Stevenson HRT and cardiovascular disease revisited

Thiolases catalyze the condensation of acyl-CoA thioesters through the Claisen condensation reaction. The best described enzymes usually yield linear condensation products. Using a combined computational/experimental approach, and guided by structural information, we have studied the potential of thiolases to synthesize branched compounds. We have identified a bulky residue located at the active site that blocks proper accommodation of substrates longer than acetyl-CoA. Amino acid replacements at such a position exert effects on the activity and product selectivity of the enzymes that are highly dependent on a protein scaffold. Among the set of five thiolases studied, Erg10 thiolase from Saccharomyces cerevisiae showed no acetyl-CoA/butyryl-CoA branched condensation activity, but variants at position F293 resulted the most active and selective biocatalysts for this reaction. This is the first time that a thiolase has been engineered to synthesize branched compounds. These novel enzy...

In this study the prevalence of myopia and age at onset among medical students were determined. Of the 140 senior medical students at The Faculty of Medicine, University of Trondheim, Norway, 133 (75 females, 58 males) were examined. Visual acuity was tested and the refractive error was measured using automated refraction and clinical refractive technique. The prevalence of myopia was found to be 50.3% in the right eye (n= 67) without significant difference between female and male students. The refractive state was unrelated to body height. Among the myopic students, the mean equivalent sphere was-2.34 f 2.01 D in the right eye (range-9.25D to-0.25 D). A clear relationship was detected between the current amount of myopia and the age at which corrective lenses were first prescribed. However, as much as 43.3% of the myopic students wearing corrective lenses first received these at the age of about 20 years, indicating a relatively high prevalence rate of adult-onset myopia.

We evaluated the impact of night reading on daytime sleepiness, quality of sleep and performance of student. Pharmacy students (N = 253, 71.5% response rate) completed a sleep and study lifestyle questionnaire, the Pittsburgh sleep quality index and the Epworth sleepiness scale questionnaires, and their grade indices (were obtained. The results showed that the prevalence of night reading was 76.7% and there was a significant (P < 0.05) association between night reading and sleep quality, and also between sleep quality and academic performance. Thus, habitual night reading may affect academic performance, possibly due to the reduced sleep and poor sleep quality of habitual night readers.

Gastric cancer remains the second leading cause of cancer deaths worldwide. Patients usually present late with local invasion or metastatic diseases. The present study investigated the expression level of liver-intestine cadherin (LI-cadherin) by RT-PCR and its correlation with clinicopathological data in 71 pairs of tumor and non-cancerous gastric mucosa. Protein expression level of LI-cadherin was determined by Western blotting and

Calcitonin gene-related peptide (CGRP), regarded by several authors to be involved in maintenance of the acetylcholine receptor, is present in the motor axons of various striated rat muscles. It is present, however, only in motor endplates of several selected striated muscles, where it is located in presynaptic axon terminals of neuromuscular junctions. No immunoreactivity could be seen within synaptic vesicles themselves. In the non-human primate Macaca fasciculata, neuromuscular junctions, including those in the diaphragm, display an intense CGRP reaction. The structure of the simian motor endplates is more elaborate than that of the rat. Amphibian motor nerve endings, both in tetanic and tonic muscles, display CGRP immunoreactivity. In tetanic m d e s the CGRP reaction outlines "terminaisons en placque" (true motor end plates) and weakly reacting "terminaisons en grappe" (grape-like endings) in tonic muscles. On supramaximal stimulation of the motor nerve, CGRP is depleted from the affected neuromuscular junctions. Wallerian degeneration of the motor axon results in complete disappearance of CGW. In most rat muscles in which motor endplates do not normally exhibit CGRP

The utilization of glycerol by Candida utilis has been studied. It has been found that this yeast has a permeability for glycerol and other three carbon compounds much greater than that of baker's yeast. This permeability allows the entrance of glycerol in Candida cells rapidly enough to permit its efficient utilization even at low concentrations. The inducibility of glycerol kinase has been established. An increase in the concentration of the mitochondria1 L-a-glycerophosphate oxidase when the yeast is grown on glycerol has also been observed. A model is presented for the substrate specificity pattern of glycerol kinase of C. mycoderma. It postulates the involvement of three hydroxyl groups in the spatial distribution corresponding to the formation of L-a-glycerophosphate from glycerol. This requirement can be met by aldoand ketotrioses in their respective hydrated forms. The pathway of glycerol formation in S. cerevisim-has also been studied. Evidence is shown of the existence of a NADH dependent enzymatic activity reducing triose phosphate to oc-glycerophosphate which can roughly account for the glycerol production. A low ionic strength seems to be required for the activity of this enzyme. The a-glycerophosphatase is specific for the L form, the efficiency of a-glycerophosphatase on D-a-glycerophosphate being 1/30 of that on L-a-glycerophosphate. The concentration of the a-glycerophosphatase in yeasts is higher when grown on hexoses than when grown on non-sugar carbon sources.

Gold nanoparticles (GNPs) offer a great promise in biomedicine. Currently, there is no data available regarding the accumulation of nanoparticles in vivo after repeated administration. The purpose of the present study was to evaluate the bioaccumulation and toxic effects of different doses (40, 200, and 400 lg/kg/day) of 12.5 nm GNPs upon intraperitoneal administration in mice every day for 8 days. The gold levels in blood did not increase with the dose administered, whereas in all the organs examined there was a proportional increase on gold, indicating efficient tissue uptake. Although brain was the organ containing the lowest quantity of injected GNPs, our data suggest that GNPs are able to cross the blood-brain barrier and accumulate in the neural tissue. Importantly, no evidence of toxicity was observed in any of the diverse studies performed, including survival, behavior, animal weight, organ morphology, blood biochemistry and tissue histology. The results indicate that tissue accumulation pattern of GNPs depend on the doses administered and the accumulation of the particles does not produce subacute physiological damage.

There is discrepancy and failure to adhere to current international guidelines for the management of metastatic colorectal cancer (CRC) in hospitals in Greece and Cyprus. The aim of the present document is to provide a consensus on the multidisciplinary management of metastastic CRC, considering both special characteristics of our Healthcare System and international guidelines. Following discussion and online communication among the members of an executive team chosen by the Hellenic Society of Medical Oncology (HeSMO), a consensus for metastastic CRC disease was developed. Statements were subjected to the Delphi methodology on two voting rounds by invited multidisciplinary international experts on CRC. Statements reaching level of agreement by ≥80% were considered as having achieved large consensus, whereas statements reaching 60-80% moderate consensus. One hundred and nine statements were developed. Ninety experts voted for those statements. The median rate of abstain per statemen...

Degenerate primers were designed from the conserved zone of hydA structural gene encoding for catalytic subunit of [Fe]-hydrogenase of different hydrogen producing bacteria. A 750 bp of PCR product was amplified by using the above-mentioned degenerate primers and genomic DNA of Enterobacter cloacae IIT-BT 08 as template. The amplified PCR product was cloned and sequenced. The sequence showed the presence of an ORF of 450 bp with significant similarity (40%) with C-terminal end of the conserved zone (H-cluster) of [Fe]-hydrogenase. hydA ORF was then amplified and cloned in-frame with GST in pGEX4T-1 and overexpressed in a non-hydrogen producing Escherichia coli BL-21 to produce a GST-fusion protein of a calculated molecular mass of about 42.1 kDa. Recombinant protein was purified and specifically recognized by anti-GST monoclonal antibody through Western blot. Southern hybridization confirmed the presence of this gene in E. cloacae IIT-BT 08 genome. In vitro hydrogenase assay with the overexpressed hydrogenase enzyme showed that it is catalytically active upon anaerobic adaptation. In vivo hydrogenase assay confirmed the presence of H 2 gas in the gas mixture obtained from the batch culture of recombinant E. coli BL-21. A tentative molecular mechanism has been proposed about the transfer of electron from electron donor to H-cluster without the mediation of the F-cluster.

BACKGROUND The use of salivary diagnostics is increasing because of its noninvasiveness, ease of sampling, and the relatively low risk of contracting infectious organisms. Saliva has been used as a biological fluid to identify and validate RNA targets in head and neck cancer patients. The goal of this study was to develop a robust, easy, and cost-effective method for isolating high yields of total RNA from saliva for downstream expression studies. METHODS Oral whole saliva (200 μL) was collected from healthy controls (n = 6) and from patients with head and neck cancer (n = 8). The method developed in-house used QIAzol lysis reagent (Qiagen) to extract RNA from saliva (both cell-free supernatants and cell pellets), followed by isopropyl alcohol precipitation, cDNA synthesis, and real-time PCR analyses for the genes encoding β-actin (“housekeeping” gene) and histatin (a salivary gland–specific gene). RESULTS The in-house QIAzol lysis reagent produced a high yield of total RNA (0.89–7....

Scorpion venom components have multifaceted orientation against bacterial, viral, fungal infections and other neuronal disorders. They can modulate the ion channels (K+, Na+, Cl−, Ca2+) of our body and this concept has been hypothesized in formulating pharmaceuticals. The triumphant achievement of these venom components as formulated anticancer agent in Phase I and Phase II clinical trials allure researchers to excavate beneficial venom components prohibiting DNA replication in malignant tumor cells. This review brings forth the achievements of Science and Technology in classifying the venom components as therapeutics and further application in drug product development.

The tightly controlled replication of hepatocytes in liver regeneration and uncontrolled proliferation of tumor cells in hepatocellular carcinoma (HCC) are often modulated by common regulatory pathways. Several microRNAs (miRNAs) are involved in HCC progression by modulating posttranscriptional expression of multiple target genes. miR-221, which is frequently up-regulated in HCCs, delays fulminant liver failure in mice by inhibiting apoptosis, indicating a pleiotropic role of miR-221 in hepatocytes. Here, we hypothesize that modulation of miR-221 targets in primary hepatocytes enhances proliferation, providing novel clues for enhanced liver regeneration. We demonstrate that miR-221 enhances proliferation of in vitro cultivated primary hepatocytes. Furthermore, applying two-thirds partial hepatectomy as a surgically induced liver regeneration model we show that adeno-associated virus-mediated overexpression of miR-221 in the mouse liver also accelerates hepatocyte proliferation in vivo. miR-221 overexpression leads to rapid S-phase entry of hepatocytes during liver regeneration. In addition to the known targets p27 and p57, we identify Aryl hydrocarbon nuclear translocator (Arnt) messenger RNA (mRNA) as a novel target of miR-221, which contributes to the pro-proliferative activity of miR-221. Conclusion: miR-221 overexpression accelerates hepatocyte proliferation. Pharmacological intervention targeting miR-221 may thus be therapeutically beneficial in liver failure by preventing apoptosis and by inducing liver regeneration. (HEPATOLOGY 2013;57:299-310) Abbreviations: AAV8, adeno-associated virus serotype-8; Arnt, aryl hydrocarbon receptor nuclear translocator; HCC, hepatocellular carcinoma; miRNA, microRNA; PTEN, phosphatase and tensin homolog; siRNA, small interfering RNA; TIMP3, tissue inhibitor of metalloproteinases 3; TTR, transthyretin; UTR, untranslated region.

For accurate and reliable gene expression results, normalization of real-time PCR data is required against a control gene, which displays highly uniform expression in living organisms during various phases of development and under different environmental conditions. We assessed the gene expression of 10 frequently used housekeeping genes, including 18S rRNA, 25S rRNA, UBC, UBQ5, UBQ10, ACT11, GAPDH, eEF-1a, eIF-4a, and b-TUB, in a diverse set of 25 rice samples. Their expression varied considerably in different tissue samples analyzed. The expression of UBQ5 and eEF-1a was most stable across all the tissue samples examined. However, 18S and 25S rRNA exhibited most stable expression in plants grown under various environmental conditions. Also, a set of two genes was found to be better as control for normalization of the data. The expression of these genes (with more uniform expression) can be used for normalization of real-time PCR results for gene expression studies in a wide variety of samples in rice.

Steady-state kinetic analyses were performed on the non-phosphorylated, in vitro phosphorylated and phosphorylation-site mutant (Ser8→Asp) forms of purified recombinant sorghum C 4 phosphoenolpyruvate (P-pyruvate) carboxylase (EC 4.1.1.3 1) containing an intact N-terminus. Significant differences in certain kinetic parameters were observed between these three enzyme forms when activity was assayed at a suboptimal but near-physiological pH (7.3), but not at optimal pH (8.0). Most notably, at pH 7.3 the apparent K i for the negative allosteric effector l-malate was 0.17 mM, 1.2 mM and 0.45 mM while the apparent K a for the positive allosteric effector glucose 6-phosphate (Glc6P) at 1mM P-pyruvate was 1.3 mM, 0.28 mM and 0.45 mM for the dephosphorylated, phosphorylated and mutant forms of the enzyme, respectively. These and related kinetic analyses at pH 7.3 show that phosphorylation of C 4 P-pyruvate carboxylase near its N-terminus has a relatively minor effect on V and K m (total P-pyruvate) but has a dramatic effect on the extent of activation by Glc6P, type of inhibition by l-malate and, most especially, K a (Glc6P) and K i (l-malate). Thus, regulatory phosphorylation profoundly influences the interactive allosteric properties of this cytosolic C 4 -photosynthesis enzyme.

There are now known to exist seven phosphoinositides all derived through various metabolic routes from the parent lipid phosphatidylinositol. With one additional metabolite, diacylglycerol, these represent a rich resource of bioactive lipids responsible for recruiting protein effectors and marking membrane compartments. The metabolic map of this pathway and the nature of the binding partner interactions are reviewed.

Objective: In this study we developed a 96-micro plate enzymatic assay for D-lactate in plasma and urine. Methods: D-lactate was assayed enzymatically with a UV-spectrophotometer in plasma from 38 and in urine from 37 diabetics and from 42 non-diabetic controls. Results: The within-run coefficients of variations (CV) were 2.6% for plasma and 5.7% for urine. The between run CVs were 6.8% for plasma and 6.7% for urine. The mean recovery with standard deviation (S.D.) was 107.4 ± 7.3% for plasma and 100.1% ± 6.7% for urine. The plasma D-lactate in diabetics were (mean ± S.D.) 39.6 ± 23.7 μM. We found significant difference between the urinary D-lactate in controls and diabetics (18.2 ± 12.0 vs. 35.9 ± 24.2 μM/mM creatinine, p b 0.0001). Conclusions: The assay proved reliable with acceptable precision and recovery. Results suggest that diabetics have elevated urinary and plasma D-lactate as compared to controls.

Increased production of amyloid β-peptide (Aβ) and altered processing of tau in Alzheimer&#39;s disease (AD) are associated with synaptic dysfunction, neuronal death and cognitive and behavioural deficits. Neuroinflammation is also a prominent feature of AD brain and considerable evidence indicates that inflammatory events play a significant role in modulating the progression of AD. The role of microglia in AD inflammation has long been acknowledged. Substantial evidence now demonstrates that astrocyte-mediated inflammatory responses also influence pathology development, synapse health and neurodegeneration in AD. Several anti-inflammatory therapies targeting astrocytes show significant benefit in models of disease, particularly with respect to tau-associated neurodegeneration. However, the effectiveness of these approaches is complex, since modulating inflammatory pathways often has opposing effects on the development of tau and amyloid pathology, and is dependent on the precise ph...

Nightshift is a common work schedule in health environments, and is associated with decreased alertness and increased adverse events at work. This decrease in alertness can be predicted from biological models of sleep homeostasis and circadian influences. Naps can provide a short-term alleviation of sleep need, and the benefits of naps have been demonstrated in laboratory-based studies, and in specific controlled work environments. The efficacy of brief naps has not been demonstrated in health workers in their usual work environment. The current study examined the effects of a 30min nap break during the nightshift in a cohort of nursing staff and scientists in their usual work environment. Measures of both subjective sleepiness and objective alertness were taken at hourly intervals throughout the nightshift, on nights where a scheduled nap was taken and nights where there was no nap. Following a nap, psychomotor performance metrics (response speed and fastest 10% reactions times) improved and self-reported sleepiness was reduced compared to nights without a nap. These improvements persisted to the end of the nightshift. Effect size estimates suggested that 20-50% of variance in these measures was explained by the interaction of time on shift and the nap intervention. Differences were found in the timing of increases in subjective and objective alertness after the nap. These findings support the effectiveness of a scheduled nap break during a nightshift to maintain alertness in health workers.

Diagnosis of autoimmune hepatitis (AIH) may be challenging. However, early diagnosis is important because immunosuppression is life-saving. Diagnostic criteria of the International Autoimmune Hepatitis Group (IAIHG) were complex and purely meant for scientific purposes. This study of the IAIHG aims to define simplified diagnostic criteria for routine clinical practice. Candidate criteria included sex, age, autoantibodies, immunoglobulins, absence of viral hepatitis, and histology. The training set included 250 AIH patients and 193 controls from 11 centers worldwide. Scores were built from variables showing predictive ability in univariate analysis. Diagnostic value of each score was assessed by the area under the receiver operating characteristic (ROC) curve. The best score was validated using data of an additional 109 AIH patients and 284 controls. This score included autoantibodies, immunoglobulin G, histology, and exclusion of viral hepatitis. The area under the curve for prediction of AIH was 0.946 in the training set and 0.91 in the validation set. Based on the ROC curves, two cutoff points were chosen. The score was found to have 88% sensitivity and 97% specificity (cutoff >6) and 81% sensitivity and 99% specificity (cutoff >7) in the validation set. Conclusion: A reliable diagnosis of AIH can be made using a very simple diagnostic score. We propose the diagnosis of probable AIH at a cutoff point greater than 6 points and definite AIH 7 points or higher. (HEPATOLOGY 2008;48:169-176.) See Editorial on Page 10 A utoimmune hepatitis (AIH) is an inflammatory condition of the liver that can affect patients of all ages, sexes, and races. 1 The diagnosis needs to be considered in any patient with elevated aminotrans-ferases. Timely diagnosis and immunosuppressive therapy contain disease activity in almost all affected patients, and various case series have reported near normal or normal life expectancy in patients diagnosed and treated adequately. 2-4 Untreated AIH, however, has a 5-year mortality above 50%. 3,5 Early diagnosis may be difficult because the clinical picture is heterogeneous and there is

Objective and rationale: Reference intervals provided on laboratory reports are essential for appropriate interpretation of test results, and can significantly impact clinical decision-making and the quality of patient care. Careful determination and/or validation of reference intervals by the laboratory for use in the patient population it serves are therefore important to ensure their proper utility. Unfortunately, critical gaps currently exist in accurate and up-to-date pediatric reference intervals for accurate interpretation of laboratory tests performed in children and adolescents. These critical gaps in the available pediatric laboratory reference intervals have the clear potential of contributing to erroneous diagnosis or misdiagnosis of many diseases of childhood and adolescence. Most of the available "normal" ranges for laboratory tests were determined over 2 decades ago on older instruments and technologies, and are no longer relevant considering the current testing technology used by clinical laboratories. It is thus critical and of utmost urgency that a more acceptable and comprehensive database be established. Discussion and conclusion: In the present review, we discuss the considerations and challenges faced when generating and validating reference intervals in accordance to the current guidelines published by the Clinical Laboratory Standards Institute (CLSI). We raise particular attention to the present-day deficiencies in available pediatric reference intervals, and highlight the special issues and unique difficulties that are additionally faced when establishing reference intervals in children. Finally, we highlight a recent Canadian initiative, the CALIPER project, whose mandate is to establish and maintain a database of comprehensive and up-to-date pediatric reference intervals to be eventually made available to all clinical laboratories worldwide.

Point your SmartPhone at the code above. If you have a QR code reader, the video abstract will appear. Or use: https://youtu.be/xwe3ZzUfGhs Background: Maternal under-nutrition due to macro and micronutrient deficiencies is a challenging public health problem worldwide, disproportionately affecting women in lowincome nations. Hence, this study aimed at assessing factors associated with dietary practice and nutritional status of pregnant women in Illu Aba Bor Zone, Southwest Ethiopia. Methods: A community-based cross-sectional study was conducted among 403 randomly selected pregnant women, from May to June, 2019. Socio-demographic and socioeconomic data were collected using a structured interviewer-administered questionnaire. Dietary Diversity was obtained using a qualitative 24-hr dietary recall method and nutritional status was assessed using mid-upper arm circumference (MUAC) measurements. Data were entered onto Epidata version 3.5.3 and exported to SPSS version 20 for analysis. Binary and multivariable logistic regression analyses were used to identify independent predictors of nutritional status and dietary practice. Odds ratio with 95% Confidence interval was estimated to identify the predictors of the outcome variables. Statistical significance was affirmed at a p-value ≤0.05. Results: The prevalence of under-nutrition among the pregnant women was 17.4% and the mean (±SD) dietary diversity score was 4.88 ± 1.27. Family-size (AOR=3, 95% CI: (1.18, 7.21)), wealth status (AOR=3.65 (1.02, 12.99)), household food security (AOR=2.59 (1.21, 5.63)) and birth interval (AOR=2.3 (1.10, 4.87)) were significantly associated with nutritional status of pregnant women. Educational status (AOR = 3.49, 95% (1.53, 7.99)), household food security (AOR = 1.61, 95% (1.02, 2.56)), nutrition knowledge (AOR = 1.74, 95% (1.08, 2.79)), and meal frequency (AOR = 2.88, 95% (1.65, 5.03)) were significantly associated with dietary practice of the respondents. Conclusion: Finding of the study revealed that the prevalence of under-nutrition and poor dietary practice was high which were significantly associated with educational and economic status. Hence, it is imperative to improve the nutritional status of pregnant women, strengthening sustainable income-generating activities and saving strategies.

Background & Aims: Therapy with interferon alfa (IFNor death. Small follow-up studies 10,11 have reported that a) leads to remission of disease in one third of patients remissions in chronic hepatitis B induced by IFN-a are with chronic hepatitis B. The aim of this study was to of long duration, but the survival of treated patients has better define the long-term prognosis of this outcome. not been well defined. A recent study reported that the Methods: One hundred three patients with chronic hepfrequency of death, liver transplantation, and severe comatitis B who underwent IFN-a therapy in three clinical plications due to cirrhosis was significantly lower among trials between 1984 and 1991 were followed up for patients who responded to IFN-a treatment than those serological status, biochemical evidence of liver diswho did not. 12 ease, and liver complications or mortality through The aim of the current study was to further evaluate 1994. Results: Among 103 patients, 31 (30%) rethe long-term survival and clinical outcome of a cohort sponded to therapy with loss of hepatitis B e antigen of patients with chronic hepatitis B who were treated and viral DNA from serum. Responders were more likely with IFN-a. than nonresponders to be women, black, and to have more severe liver disease including cirrhosis (P õ Materials and Methods 0.05

Advancements in the quality and availability of highly sensitive analytical instrumentation and methodologies have led to increased interest in the use of microsamples. Among microsamples, dried blood spots (DBS) are the most well-known. Although there have been a variety of review papers published on DBS, there has been no attempt at describing the full range of analytes measurable in DBS, or any systematic approach published for characterizing the strengths and weaknesses associated with adoption of DBS analyses. A scoping review of reviews methodology was used for characterizing the state of the science in DBS. We identified 2018 analytes measured in DBS and found every common analytic method applied to traditional liquid samples had been applied to DBS samples. Analytes covered a broad range of biomarkers that included genes, transcripts, proteins, and metabolites. Strengths of DBS enable its application in most clinical and laboratory settings, and the removal of phlebotomy and...

Objectives: To compare four methods for thyroglobulin (Tg) quantitation and three methods for detection and quantitation of thyroglobulin antibodies (Tg-Ab). We also wanted to explore the premise that thyroglobulin antibodies, as determined by commercially available assays, interfere with thyroglobulin assays in a predictable way. Methods: Split sample method comparisons were run on all the methods for both the thyroglobulin and anti-thyroglobulin assays. In addition to this, samples from patients that had disseminated thyroid cancer but had low serum thyroglobulin concentrations and high thyroglobulin antibodies were further studied. These studies involved doing recovery studies (or antibody inhibition studies). Results: There was good agreement between methods for quantitation of thyroglobulin with slopes ranging from 0.77 to 1.23 although closer agreement was expected as the assays are all calibrated to the same reference standard (CRM 457). The situation for the thyroglobulin antibody assays is significantly worse, and the rate of antibody positivity ranged from 9-21% in this group of patients although there was agreement in only 6%. Different reference standards are used for the Tg-Ab assays we investigated. The Tg-Ab data did not lend itself to traditional linear regression analysis as the data showed wide scatter. Conclusions: There is good agreement between the four thyroglobulin assays compared in this study. The linear regression analysis shows that there is proportional error present between the methods that is greater than 50%. This study is unable to demonstrate any difference in assay values based on the amount of anti-thyrogobulin present in the specimen. The agreement between different anti-thyroglobulin assays is very poor. This finding is very problematic since it makes it difficult to generalize any literature reports of interference. All the thyroglobulin assays appear to be suitable for monitoring patients with thyroid cancer, provided that the differences in calibration are taken into account. Differences in calibration between different assays need to be taken into account when changing assays. Conversely, the anti-thyroglobulin assays are virtually useless since there appears to be very little agreement between the three assays studied and no evidence of assay interference in the measurements of thyroglobulin.

Insulin-like peptide 5 (INSL5) is an orexigenic peptide hormone belonging to the relaxin family of peptides. It is expressed primarily in the L-cells of the colon and has a postulated key role in regulating food intake. Its G protein-coupled receptor, RXFP4, is a potential drug target for treating obesity and anorexia. We studied the effect of modification of the C-terminus of the A and B-chains of human INSL5 on RXFP4 binding and activation. Three variants of human INSL5 were prepared using solid phase peptide synthesis and subsequent sequential regioselective disulfide bond formation. The peptides were synthesized as C-terminal acids (both A- and B-chains with free C-termini, i.e., the native form), amides (both chains as the C-terminal amide) and one analog with the C-terminus of its A-chain as the amide and the C-terminus of the B-chain as the acid. The results showed that C-terminus of the B-chain is more important than that of the A-chain for RXFP4 binding and activity. Amidat...

Cyanobacterial species composition of fresh water and terrestrial ecosystems and chemical environment of water in Schirmacher Oasis in Continental Antarctica was investigated. Over 35 species of cyanobacteria were recorded. Diazotrophic species both heterocystous and unicellular contributed more than half to the count except in lake ecosystem. The species composition varied among the fresh water as well as terrestrial ecosystems. The physico-chemical analyses of water revealed its poor nurient content which might have supported the growth of diazotrophic cyanobacteria in an Antarctic environment. Among the cyanobacteria Oscillatoria, Phormidium and Nostoc commune were the dominant flora in most of the habitats. The physiological characteristics of isolated cyanobacteria strains indicated that N2-fixation, nitrate uptake, nitrate-reduction, ammonium-uptake, GS-transferase activity and photosynthesis was unaffected at low temperature (5 degrees C) which indicated low temperature adapt...

Parkinson's disease is the second most common neurodegenerative disease, after Alzheimer's disease, among the aging human population. The main symptoms of Parkinson's disease such as tremor and movement disabilities are the result of degeneration of dopaminergic neurons in substantia nigra pars compacta. The widely-accepted subcellular factor which underlies Parkinson's disease neuropathology is the presence of Lewy bodies with characteristic inclusions of aggregated alpha-synuclein. This small soluble protein has been implicated in a range of interactions with phospholipid membranes and free fatty acids. The precise biological function of this protein is, however, still under investigation. Here we review the evidence linking alpha-synuclein, lipid metabolism, fatty acid oxidation, mitochondrial damage and Parkinson's disease. We propose that association of alpha-synuclein with oxidized lipid metabolites can lead to mitochondrial dysfunction in turn leading to dopaminergic neuron death and thus to Parkinson's disease.

Native Paralithodes camtschaticae hemocyanin is found as a mixture of dodecamers (24S; 80%) and hexamers (16S; 20%). Removal of Ca 21 ions by dialysis against EDTA-containing buffer solution at neutral pH induces complete dissociation of the 24S form into the 16S form. Under these conditions, a further increase in pH to 9.2 produces complete dissociation of the hexamers into monomers (5S). In both cases, the dissociation process is reversible. The dodecamer (24S) is composed of two different hexamers which can be discriminated only by ion-exchange chromatography in the presence of Ca 21 ions. At alkaline pH and in the presence of EDTA, two major monomeric fractions can be separated by ion-exchange chromatography: ParcI (60%) and ParcII (40%). The reassociation properties of the two fractions were studied separately to define their ability to form hexamers and dodecamers.

The pigment melanin, which is believed to play a photoprotective role, was quantified here in human RPE cells from donors of different age. Electron spin resonance (ESR) spectroscopy was shown to provide a quantitative measure of melanin and was used as a non-destructive measure of melanin content. Results indicated an age-related melanin loss in RPE cells, with melanin content diminishing 2·5-fold between the first and the ninth decade of life. To determine whether photo-oxidation may contribute to age-related changes in RPE melanin, RPE in human eyecups, isolated human and bovine RPE cells, purified melanin granules, or synthetic dopa melanin were irradiated with various wavelengths and intensities of visible light. Samples were analysed for changes in melanin content by ESR spectroscopy, and by absorption and emission spectrophotometry. The concentration of hydrogen peroxide was measured in some samples, and some human eyecups were examined by transmission electron microscopy. Irradiation of RPE in eyecups with intense visible light was found to produce a timedependent photobleaching of melanosomes that was accompanied by the formation of hydrogen peroxide. Photobleaching of isolated RPE melanosomes and synthetic dopa melanin resulted in enhanced melanin fluorescence, as previously shown for melanin from aged donors by others, and significantly reduced ESR signal intensity, resembling the changes in melanin with aging observed here. We conclude that the content of melanin in RPE cells undergoes an age-related change to which photo-oxidation may contribute. This observation raises the question of whether age-related changes in melanin reduce the photoprotective role of the pigment in aging RPE cells.

Neural invasion by pancreatic cancer cells (PCC) worsens the prognosis and frequently limits curative resection. We established a novel in-vitro model in which T3M4-PCCs were co-cultured with either isolated myenteric plexus cells (MP) or dorsal root ganglia (DRG) of newborn rats within a three-dimensional extracellular matrix gel. The close vicinity of MP or DRG to T3M4-PCCs induced early morphologic changes on T3M4-PCCs at the migration front prior to the migration process with elongated and neurite-targeting PCCs, compared to round and non-grouping at the non-migrating front. T3M4-PCCs built cancer-cell clusters around the DRG or MP, a process which was accelerated by increasing number of T3M4-PCCs or neurons. These findings indicate that neuro-cancer interactions start prior to PCC migration and induce evident changes in cancer and nerve biology. These findings can be reproduced within the introduced 3D in-vitro migration assay which allows investigation in the early pathogenesis of neural PCC invasion.

The genus Achillea genus houses more than 100 species, a number of them are popularly used in traditional medicine for spasmodic gastrointestinal, gynecological and hepatobiliary disorders, hemorrhages, pneumonia, rheumatic pain, inflammation, wounds healing etc. Members of the genus contain a wide variety of volatile and non-volatile secondary metabolites, including terpenes, polyphenols, flavonoids and others. Multiple studies have assessed the biological effects and other aspects of Achillea spp. In a number of preclinical studies, Achillea plants and their essential oils have demonstrated promising antibacterial properties against a number of human and plant pathogens. Besides, the plants have displayed strong antioxidative and potent anti-proliferative and anticancer properties in various cellular and animal models. Achillea plants have widely been used as food preservative in food industry. Clinical studies have indicated its potential against multiple sclerosis (MS), irritable bowel syndrome (IBS), ulcerative colitis, episiotomy wound, primary dysmenorrhea, oral mucositis etc. The present work focuses to provide a brief overview on folk knowledge, phytochemistry, biological activity and applications of Achillea plants. There is a close relationship between the traditional ethnobotanical usage and pharmacological and clinical data from different Achillea spp. The application of Achillea plants and their extracts seems to be a promising alternative for antimicrobial and antioxidant purposes in food, pharmaceutical and cosmetic industries.

This review focuses on recent insights into the mechanisms and the biological functions of the proteasome. This large ATP-dependent proteolytic complex is the main site for protein degradation in mammalian cells and catalyses the rapid degradation of ubiquitinated proteins, and is the source of most antigenic peptides used by the immune system to screen for viruses and cancer. ATP is required to unfold globular proteins to open the gated channel into the 20S proteasome and to facilitate protein translation into it. Inhibitors of its proteolytic activity are widely used as research tools and have proven effective in cancer therapy.

Fatigue associated with cholestasis may impair health-related quality of life. The pathogenesis of this symptom is largely unknown, but it has been suggested that central serotoninergic neurotransmission may be implicated and that serotonin 1A receptor agonists may yield improvement. The aim of this study was to study the central serotoninergic system, specifically the serotonin (5-HT) 1A receptor-mediated pathway of serotoninergic neurotransmission, in a bile duct resection rat model of cholestasis. Fatigue was assessed in the forced swim test in sham and bile duct-resected rats. The serotonin behavioral syndrome, which includes hyperlocomotion, was assessed in both groups of rats after escalating doses of the 5-HT 1A receptor agonist 8-hydroxy(di-n-propylamine)tetralin (8-OH DPAT). 5-HT 1A and 5-HT 2 receptor densities were explored in four brain regions using a receptor-binding assay. Extracellular 5-HT and 5-hydroxyindoleacetic acid were measured via in vivo brain dialysis. Bile duct-resected rats spent more time floating in the forced swim test, and 8-OH DPAT decreased floating time in cholestatic rats (P < .01). Dose-response curves created with 8-OH DPAT for the serotonin behavioral syndrome were similar in bile duct-resected and sham-operated rats. 5-HT 1A and 5-HT 2 receptor densities in most brain regions and extracellular serotonin levels were similar in both groups of rats. In conclusion, 5-HT 1A receptor agonist-induced amelioration of fatigue in cholestatic rats may be nonspecific and not linked to reversal of the pathophysiology of fatigue associated with cholestasis; however, these data do not exclude a potential role of the central serotoninergic system in the evolution of fatigue. (HEPATOLOGY 2005;41:731-737.

This morphology imposed the concept of a soluble transmitter, released or modulated at the level of the disc membrane and acting at the level of the cellular membrane. Early studies had concentrated on the photochemistry of retinal, the molecule acted upon by light, which induces its isomerization from the 11-cis to the all-trans conformation. As this transconformation is followed by the release of the all-trans retinal from its membrane-bound protein cofactor This review is dedicated to thc memory of our friend Hermann Kiihn, a pioneer of the biochemistry of visual transduction.

Objectives: Procalcitonin (PCT) and interleukin-6 (IL-6) are established markers of tissue inflammation and injury. The aim of the present study was to investigate the possible correlation of PCT and IL-6 with liver metastasis. Design and methods: The study consisted of fifteen healthy controls (group A), twenty-one patients with solid tumors without metastases (group B), eleven patients with liver metastasis only (group C) and eleven patients with generalized metastatic disease (group D). Results: Serum PCT levels were significantly increased in group D compared to groups A (p < 0.001) and B (p = 0.004), but no difference was observed in PCT levels between groups C and B or C and D. IL-6 serum levels were markedly elevated in group C compared to group A (p < 0.001) or to groups B (p < 0.001) and D (p = 0.02). A positive correlation was observed between PCT and IL-6 serum levels (r = 0.357, p = 0.019). Conclusions: PCT levels are related to disease stage in cancer patients, whereas IL-6 concentration seems to be a more specific marker of liver metastasis.

Inhibition of fatty acid synthase (FAS) induces apoptosis in human breast cancer cells in vitro and in vivo without toxicity to proliferating normal cells. We have previously shown that FAS inhibition causes a rapid increase in malonyl-CoA levels identifying malonyl-CoA as a potential trigger of apoptosis. In this study we further investigated the role of malonyl-CoA during FAS inhibition. We have found that: [i] inhibition of FAS with cerulenin causes carnitine palmitoyltransferase-1 (CPT-1) inhibition and fatty acid oxidation inhibition in MCF-7 human breast cancer cells likely mediated by elevation of malonyl-CoA; [ii] cerulenin cytotoxicity is due to the nonphysiological state of increased malonyl-CoA, decreased fatty acid oxidation, and decreased fatty acid synthesis; and [iii] the cytotoxic effect of cerulenin can be mimicked by simultaneous inhibition of CPT-1, with etomoxir, and fatty acid synthesis with TOFA, an acetyl-CoA carboxylase (ACC) inhibitor. This study identifies CPT-1 and ACC as two new potential targets for cancer chemotherapy.

The studies of the cytochrome P450 (P450) arachidonic acid (AA) monooxygenase, now established as a major pathway for the bioactivation of AA, have uncovered new and important functional roles for this enzyme system in cell and organ physiology, and in the metabolism of endogenous substrate. Past and present advances in P450 biochemistry and molecular biology are beginning to provide a description of the P450 isoform specificity of AA bioactivation, and the mechanisms of action and physiological relevance of the P450 metabolites. Associations between genetically controlled alterations in P450 function, expression, or regulation and functionally meaningful phenotypes point to the critical roles played by the AA monooxygenase in the control of systemic blood pressure and the pathophysiology of hypertension.

N-Acylethanolamines (NAEs) constitute a new class of plant lipids and are thought to play a role in plant defense strategies against pathogens. In plant defense systems, oxylipins generated by the lipoxygenase pathway are important actors. To date, it is not known whether plants also use endogeneous oxylipins derived from NAEs in their defense reactions. We tested whether members of the NAE class can be converted by enzymes constituting this pathway, such as (soybean) lipoxygenase-1, (alfalfa) hydroperoxide lyase and (flax seed) allene oxide synthase. We found that both a-N-linolenoylethanolamine and g-N-linolenoylethanolamine (18:3), as well as a-N-linolenoylamine and g-N-linolenoylamine were converted into their (13S )-hydroperoxide derivatives by lipoxygenase. Interestingly, only the hydroperoxides of a-N-linolenoyl(ethanol)amines and their linoleic acid analogs (18:2) were suitable substrates for hydroperoxide lyase. Hexanal and (3Z )-hexenal were identified as volatile products of the 18:2 and 18:3 fatty acid (ethanol)amides, respectively. 12-Oxo-N-(9Z )dodecenoyl(ethanol)amine was the nonvolatile hydrolysis product. Kinetic studies with lipoxygenase and hydroperoxide lyase revealed that the fatty acid ethanolamides were converted as readily or even better than the corresponding free fatty acids. Allene oxide synthase utilized all substrates, but was most active on (13S )-hydroperoxy-a-N-linolenoylethanolamine and the (13S )-hydroperoxide of linoleic acid and its ethanolamine derivative. a-Ketols and g-ketols were characterized as products. In addition, cyclized products, i.e. 12-oxo-N-phytodienoylamines, derived from (13S )-hydroperoxy-a-N-linolenoylamines were found. The results presented here show that, in principle, hydroperoxide NAEs can be formed in plants and subsequently converted into novel phytooxylipins.

LL-37 is an amphipathic, R-helical, antimicrobial peptide. 15 N chemical shift and 15 N dipolarshift spectroscopy of site-specifically labeled LL-37 in oriented lipid bilayers indicate that the amphipathic helix is oriented parallel to the surface of the bilayer. This surface orientation is maintained in both anionic and zwitterionic bilayers and at different temperatures and peptide concentrations, ruling out a barrelstave mechanism for bilayer disruption by LL-37. In contrast, electrostatic factors, the type of lipid, and the presence of cholesterol do affect the extent to which LL-37 perturbs the lipids in the bilayer as observed with 31 P NMR. The 31 P spectra also show that micelles or other small, rapidly tumbling membrane fragments are not formed in the presence of LL-37, excluding a detergent-like mechanism. LL-37 does increase the lamellar to inverted hexagonal phase transition temperature of both PE model lipid systems and Escherichia coli lipids, demonstrating that it induces positive curvature strain in these environments. These results support a toroidal pore mechanism of lipid bilayer disruption by LL-37.