Enzyme-linked Immune Absorbent Assay Research Articles

Comparison of Urinary Biomarkers in Diagnosis of Ureteropelvic Junction Obstruction and Differentiation from Nonobstructive Dilatation.

Differentiation of nonobstructive dilatation (NOD) from ureteropelvic junction obstruction (UPJO) is a challenge in children with antenatally detected hydronephrosis. The aim of this study is to compare the utility of urinary biomarkers: carbohydrate antigen (CA 19-9), neutrophil gelatinase-associated lipocalin (NGAL), and kidney injury molecule (KIM) in diagnosing UPJO. A prospective study was conducted after obtaining ethical clearance between 2021 and 2022. Group 1 - control group (n = 30): children with normal antenatal ultrasound with no urinary symptoms. Group 2 - study group (n = 48): children with unilateral hydronephrosis: Group 2a - NOD (n = 24): children stable on ultrasound and diuretic renogram and Group 2b - UPJO (n = 24): children who worsened to Grade 4 hydronephrosis on ultrasound/worsening of differential renal function (10% drop) on renogram who underwent pyeloplasty. Urinary biomarkers NGAL, KIM-1, and CA 19-9 were measured using the enzyme-linked immune absorbent assay method. The urine CA 19-9 level was 128.05 ± 4.08 U/mL in the UPJO group, and this was significantly higher (P = 0.001) than NOD, 70.29 ± 4.41, and controls, 1.91 ± 1.57. The urine NGAL level was 21.41 ± 4.44 pg/mL in UPJO, and this was significantly higher than controls, 2.669 ± 0.513, but not NOD, 24.55 ± 2.67. The urine KIM level was 817 ± 15.84 pg/mL in the UPJO group, and this was significantly higher than controls, 285 ± 8.10, but not NOD, 768.23 ± 15.12. Receiver operating characteristic analysis of CA 19-9 revealed a urine biomarker cutoff of 95 U/mL for diagnosing UPJO (sensitivity 95%; specificity 96%; and area under the curve 0.99). CA 19-9 is a superior marker compared to NGAL and KIM in differentiating UPJO from NOD. Further studies with larger numbers are warranted.

Parental stress parameter difference after caring for congenital adrenal hyperplasia children with genital ambiguity

Abstract Introduction: Congenital adrenal hyperplasia (CAH) disrupts cortisol biosynthesis, which may cause genital ambiguity in females. Caring for children with genital ambiguity creates parental distress. Common stress parameters include biological (e.g., serotonin cortisol) and psychological (e.g., Depression, Anxiety, and Stress Scale [DASS] scores). This study elaborated parental stress parameter differences among parents of CAH children with and without genital ambiguity. Methods: This cross-sectional study included 34 fathers and 44 mothers of children with CAH. All participants agreed to fill in the DASS-42 and socioeconomic questionnaire. Morning serum serotonin and cortisol samples were examined using the enzyme-linked immune-absorbent assay technique. Results: The median (minimum–maximum) value of serotonin concentration was lower in fathers compared to mothers (281.18 [78.44–122.91] ng/ml vs. 399.78 [122.91–1050.0] ng/ml, P = 0.11); in parents of children with genital ambiguity (292.22 [78.44–1050.00] ng/ml vs. 426.71 [150.78–1050.00] ng/ml, P = 0.01); and in parents of children with a diagnosis time <1 year (225.00 [93.41–570.33] ng/ml vs. 371.815 [78.44–1050.00] ng/ml, P = 0.046). Higher cortisol concentration was observed in parents of children with diagnosis time <1 year (78.65 [50.89–126.89] ng/ml vs. 63.33 [6.14–283.67] ng/ml, P = 0.046). The proportion of depression was 24.4%, anxiety 45.1%, and stress 21.8%. Conclusion: Fathers, parents of children with genital ambiguity, and being recently diagnosed had lower serum serotonin levels. Parents of recently diagnosed children had higher cortisol levels. Anxiety was more common than depression and stress.

Assessment of salivary and gingival crevicular fluids IL6, IL-8, IL-12, and IL-17 levels in healthy subjects and patients with periodontal disease.

Biomarkers present in the oral fluid (saliva and GCF) have been studied thoroughly in order to be used as adjuncts to the diagnosis of periodontal disease. In periodontal diseases, inflammatory mediators, including IL6, IL-8, IL-12, and IL-17, may promote the degeneration of periodontal tissues. The aim of the present study was to quantify IL6, IL-8, IL-12, and IL-17 levels in the saliva and GCF for patients with generalized periodontitis and compare it with healthy subjects. Furthermore, identifying the most reliable biofluids (saliva and GCF) that can be utilized for biomarkers detection in periodontitis patients. In this study, GCF and saliva samples were collected from 40 patients with generalized periodontitis and 20 healthy subjects. Enzyme-linked immune absorbent assay (ELISA) was used in the measurement of interleukin levels. The mean levels of IL6, IL-8, IL-12, and IL-17 either in saliva or GCF were higher in periodontitis patients than in healthy subjects. These results suggest that the levels of IL6, IL-8, IL-12, and IL-17 in GCF or saliva may be relevant in the pathophysiology of periodontitis, and the measurement of these cytokines may be beneficial in the identification of patients with periodontitis. Furthermore, some biomarkers for periodontal disease are detected better in saliva, while others are best detected in GCF.

Environmental Enrichment Protects Offspring of a Rat Model of Preeclampsia from Cognitive Decline.

Preeclampsia affects 5-7% of all pregnancies and contributes to adverse pregnancy and birth outcomes. In addition to the short-term effects of preeclampsia, preeclampsia can exert long-term adverse effects on offspring. Numerous studies have demonstrated that offspring of preeclamptic women exhibit cognitive deficits from childhood to old age. However, effective ways to improve the cognitive abilities of these offspring remain to be investigated. The aim of this study was to explore whether environmental enrichment in early life could restore the cognitive ability of the offspring of a rat model of preeclampsia and to investigate the cellular and molecular mechanisms by which EE improves cognitive ability. L-NAME was used to establish a rat model of preeclampsia. The spatial learning and memory abilities and recognition memory of 56-day-old offspring were evaluated by the Morris water maze and Novel object recognition (NOR) task. Immunofluorescence was performed to evaluate cell proliferation and apoptosis in the DG region of the hippocampus. qRT-PCR was performed to examine the expression levels of neurogenesis-associated genes, pre- and postsynaptic proteins and inflammatory cytokines. An enzyme-linked immune absorbent assay was performed to evaluate the concentration of vascular endothelial growth factor (VEGF) and inflammatory cytokines in the hippocampus. The administration of L-NAME led to increased systolic blood pressure and urine protein levels in pregnant rats. Offspring in the L-NAME group exhibited impaired spatial learning ability and memory as well as NOR memory. Hippocampal neurogenesis and synaptic plasticity were impaired in offspring from the L-NAME group. Furthermore, cell apoptosis in the hippocampus was increased in the L-NAME group. The hippocampus was skewed to a proinflammatory profile, as shown by increased inflammatory cytokine levels. EE improved the cognitive ability of offspring in the L-NAME group and resulted in increased hippocampal neurogenesis and synaptic protein expression levels and decreased apoptosis and inflammatory cytokine levels. Environmental enrichment resolves cognitive impairment in the offspring of a rat model of preeclampsia by improving hippocampal neurogenesis and synaptic plasticity and normalizing the apoptosis level and the inflammatory balance.

Potential Role of Macrophage Phenotypes and CCL2 in the Pathogenesis of Takayasu Arteritis.

ObjectivesTo investigate vascular macrophage phenotype as well as vascular and peripheral chemokine (C-C motif) ligand 2 (CCL2) expression during different stages of disease progression in patients with Takayasu Arteritis (TA).MethodsIn this study, 74 patients with TA and 50 controls were recruited. TA disease activity was evaluated with Kerr scores. Macrophage phenotype and CCL2 expression were examined by immunohistochemistry in vascular specimens from 8 untreated and 7 treated TA patients, along with 4 healthy controls. Serum CCL2 were quantified by enzyme-linked immune-absorbent assay from TA patients at baseline (n=59), at 6-months (n=38), and from 46 healthy volunteers. Vascular macrophage phenotype, vascular CCL2 expression and serum CCL2 levels during different stages, as well as the relationship between serum CCL2 and disease activity or other inflammatory parameters (erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and interleukin 6 (IL-6)) were investigated.ResultsIn untreated patients, vascular M1 macrophages and CCL2 showed increased expression, mainly in the adventitia. In contrast, in treated patients, vascular adventitial M1 and CCL2 expression were decreased, while vascular medial M2 macrophages and CCL2 levels were increased. Distribution of macrophages and CCL2 was consistent within the TA vascular lesions regardless of the disease stage. Furthermore, peripheral CCL2 was elevated in patients with TA (TA: 160.30 ± 120.05 vs. Control: 65.58 ± 54.56 pg/ml, P < 0.001). CCL2 levels were found to correlate with ESR, CRP, and IL-6 (all R values between 0.55 and 0.6, all P < 0.001). Receiver operating curve analysis demonstrated that CCL2 (at the cut-off value of 100.36 pg/ml) was able to predict disease activity (area under the curve = 0.74, P = 0.03). Decrease in CCL2 level was observed in patients with clinical remission (CR), but not in patients without CR, after 6 months of treatment (CR patients: baseline 220.18 ± 222.69 vs. post-treatment 88.71 ± 55.89 pg/ml, P = 0.04; non-CR patients: baseline 142.45 ± 104.76 vs. post-treatment 279.49 ± 229.46 pg/ml, P = 0.02).ConclusionsMacrophages contribute to vascular pathological changes in TA by undergoing phenotype transformation. CCL2 is an important factor for recruiting macrophages and a potential biomarker for disease activity.

Screening of the aflatoxin M1 metabolite in urine samples of residents in Terengganu, Malaysia

A study was conducted to screen the occurrence and level of aflatoxin M1 (AFM1) in urine samples of 206 urban and rural residents in Terengganu, Malaysia. The level of AFM1 was quantified by competitive enzyme-linked immune-absorbent assay (ELISA). Of the 206 samples, 84 were positive for AFM1 (40.8%) in a range of 0.07–5.53 ng/ml (mean = 0.589 ng/ml). Residents of Terengganu are moderately exposed to AFM1. Age, ethnicity, marital status and employment status were associated with urinary level of AFM1. Subjects aged 30 years and above, non-Malays, married, and those unemployed had significantly higher levels of urinary AFM1 (p < 0.05). Since aflatoxin is recognised as a potent-carcinogen for liver cancer and a continuous exposure to this toxin can be fatal, the present findings could provide a baseline for future studies where larger samples and more advanced techniques might be used to find the possible effects of the exposure of this toxin on the community's health.

Serum levels of retinol-binding protein 4 and the risk of non-small cell lung cancer

Retinol binding protein 4 (RBP4), as an adipokine, has been identified to be associated with several types of cancer. However, no studies have assessed its effect on non-small cell lung cancer (NSCLC) risk. The objective of this study was to assess the association between serum RBP4 levels and the risk of NSCLC.A case-control study design was used to recruit 256 confirmed NSCLC cases and 256 age- and gender-matched healthy controls by frequency between August 2017 and January 2019. Serum RBP4 was measured using enzyme-linked immune absorbent assay before treatment. Unconditional logistic regression analysis was applied to estimate the odds ratio and 95% confidence interval (CI).Serum RBP4 level was significantly higher in NSCLC patients than those in the healthy control group (36.05 ± 8.28 vs 29.54 ± 7.71 μg/mL, P < .05). Higher serum RBP4 level was associated with increased risk of NSCLC (P trend = .001). Compare with those in the lowest tertile, the adjusted odds ratios were 1.85 (95% CIs 1.07–3.2) (P = .029) for the second tertile and 2.18 (95% CIs 1.37–3.45) (P = .001) for the highest tertile after adjusting for confounding variables. No interactions were observed after stratified analyses by body mass index and smoking status (P for interaction: .584 and .357).Our study indicated that serum RBP4 level was positively related to the risk of NSCLC. Additional studies with prospective design are required to confirm this finding.

Detection of Brucellosis in Blood of Cows in Some Farms of Khartoum State

Brucellosis is an infectious disease that affect human as well as animals. The disease is caused by exposure to a bacterial species belong to the genus Brucella. Brucellosis has wide distribution in sub-Saharan African countries, including Sudan in which the diagnosis remains a great challenge and basically depend on serology. The present study was carried out on five dairy cattle farms allegedly free from brucellosis, but with sporadic cases of abortion. As well-known Sudan has a many dairy industries with peri‑urban dairy establishments which built in order to improve milk availability to rural communities. The aim of this study is to estimate the prevalence of brucellosis in Sudan, using samples submitted to collect from different farms. Five farms were tested with Rose Bengal Test (RBT) and Enzyme-linked Immune-absorbent Assay (ELISA). Parallel testing was used to determine whether a farm was to be considered positive or not. The result showed that out of 70 cow’s Blood samples examine 22 (31.4%) had positive RBPT results while the rest (68.6%) had Negative RBPT results. Of the Five farms which were examined the positive cows were observed in only 2 farms. The results show the need to carry out surveillance of brucellosis in cattle in Sudan to understand the spatial distribution of the disease in the country. These surveillance could reduce the risk of Brucellosis spreading and consequently improvement will take place in the quality of livestock and their dairy product which consumed by the human in those areas.

1,25(OH)2D3 Activates Autophagy to Protect against Oxidative Damage of INS-1 Pancreatic Beta Cells.

Diabetes mellitus is a serious disease endangering human health worldwide. Vitamin D (Vit D) is a well-characterized regulator of calcium-phosphorus metabolism that also exerts other biological effects extending far beyond mineral homeostasis. Some epidemiological studies have suggested that Vit D has a role in defense against diabetes, although the mechanism remains unclear. Autophagy, an intracellular catabolic process, is necessary to maintain the normal structure and function of host cells. In our previous study, we found that Vit D could induce autophagy of pancreatic beta cells and prevent insulitis, although the underlying mechanisms remain to be fully elucidated. In this study, the protective effect of 1,25(OH)2D3, the physiologically active metabolite of Vit D, against streptozotocin-induced cytotoxicity in rat insulinoma cell line (INS-1) cells was explored. Cell viability and insulin secretion of INS-1 cells in response to different treatments were measured with a cell counting kit and enzyme-linked immune absorbent assay (ELISA), respectively. In addition, malondialdehyde (MDA) content and total antioxidant capacity (T-AOC) were measured by ELISA. RT-PCR and Western blot analyses were used to detect autophagy levels, reactive oxygen species (ROS) was assessed by fluorescence microscope, ultrastructure analysis was performed using transmission electron microscopy. The results demonstrated that 1,25(OH)2D3 could increase cell viability and insulin secretion of INS-1 cells, and protected cells from oxidative damage induced by streptozotocin (STZ) through autophagy activation. These findings shed light on mechanisms underlying the ameliorative effects of Vit D on diabetes mellitus.

Oncogenic Wnt3a: A Candidate Specific Marker and Novel Molecular Target for Hepatocellular Carcinoma.

Background and aim: It is of the utmost importance for the specific diagnosis and effective therapy of hepatocellular carcinoma (HCC). Abnormality of oncogenic Wingless-type MMTV integration site family member 3a (Wnt3a) has been associated with progression of HCC. In this study, we aimed to evaluate Wnt3a as a novel biomarker and target for HCC.Methods: Circulating Wnt3a levels were quantitatively detected in a cohort of chronic liver diseases by an enzyme-linked immune-absorbent assay (ELISA). Hepatic Wnt3a expression in HCC and para-cancerous tissues was analyzed by immunohistochemistry. Prognostic value of Wnt3a for HCC was discovered in the cohort from the Cancer Genome Atlas (TCGA). Dynamic alterations of Wnt3a levels were detected in the hepatocarcinogenesis model induced by 2-acetylaminofluorene. Effects of Wnt3a on biological behaviors were evaluated in vitro and in vivo based on Crispr/Cas9.Results: Up-regulated Wnt3a levels were observed in serum of HCC patients with high specificity and sensitivity for HCC diagnosis. Combination of Wnt3a and AFP could improve sensitivity to 93.9% in serological detection. In addition, Wnt3a expression in HCC tissues was significantly higher than that in para-cancerous tissues. The cohort of TCGA demonstrated that high Wnt3a expression led to a poor survival of HCC patients, especially in cases at advanced stages. Furthermore, the hepatocarcinogenesis model showed that Wnt3a dynamically increased in the development of HCC. Functionally, silencing Wnt3a by Crispr/Cas9 suppressed the proliferation, colony formation, induced cell cycle arrest of HCC cells by de-activating Wnt/β-catenin pathway in vitro, and inhibited xenograft tumor growth in vivo.Conclusions: Oncogenic Wnt3a could be considered as a candidate biomarker and novel target for HCC.

Environmental enrichment reduces adolescent anxiety- and depression-like behaviors of rats subjected to infant nerve injury

BackgroundInfant nerve injury causes delayed adolescent neuropathic pain, but whether it also leads to psychiatric illness is unknown. Environmental enrichment (EE) increases social communication and activity. Thus, our goal was to test anxiety- and depression-like behaviors after infant peripheral nerve injury and evaluate the effect of environmental enrichment on these models of affective disorders.MethodsOpen field, elevated plus maze, sucrose preference, and pain behaviors (paw withdrawal threshold, spontaneous guarding score, and cold response to acetone) were measured in rats that received infant spared nerve injury (SNI). Enzyme-linked immune absorbent assay of cytokines was performed to evaluate the inflammatory response in the brain. Then, the ability of intracerebroventricular (ICV) injection of a microglia inhibitor, minocycline (MIN), and EE (a free-running wheel, a staircase, a plastic tunnel, a raised platform, and various colored balls) to reverse the infant SNI effects on behaviors and cytokines was examined.ResultsInfant nerve injury resulted in adolescent anxiety- and depression-like behaviors. The medial prefrontal cortex, basolateral amygdala, and ventral hippocampus were skewed to a pro-inflammatory profile. ICV injection of MIN reduced anxiety- and depression-like behaviors without affecting pain behaviors. In addition, ICV MIN skewed the brain towards an anti-inflammatory profile. Finally, environmental enrichment improved anxiety- and depression-like behaviors, as well as pain behaviors. EE increased brain IL-10 and decreased IL-1β and TNF-α.ConclusionsInfant nerve injury induces adolescent anxiety- and depression-like behaviors and central nervous inflammation. Environmental enrichment reduces these behaviors by normalizing the inflammation balance in the brain.

Sirtuin1, Inducible Nitric Oxide Synthase in Newly Diagnosed Iraqi Diabetic Patients

In Mammals Sirtuins (SIRTs) are members of the silent information regulator two family Sirtuin1 regulate a variety of cellular functions. It shows anti-oxidative, anti-inflammatory, and anti-apoptotic effects against cellular damage.SIRT1 conserves the cells by the mitochondrial biogenesis organizing, cellular energy and oxido reduction state. Vascular tissues are protected by SIRT1 too. In diabetic patients mutation in exon1 of SIRT1 (L107P) in insulin-generating cells conducted in over production of nitric oxide synthase (iNOS), TNF-α.Assessment of SIRT1 protein levels in newly diagnosed Iraqi diabetic in regards to family history Age, gender, and duration of disease, and its correlation with biochemical parameter in the same group. Measurement of iNOS, TNF-α.This study involved totally 40 volunteers. This group were subdivided (20) volunteers as newly diagnosis (DMT1) duration disease ˂1 year, group1 was subdivided to 10 male and 10 female: (10 with family history and 10 without family history) and group two healthy volunteers. Group1 All DMT1 were under insulin treatment. Healthy Volunteers (20) included in this study as control groups. None Healthy Volunteers were alcoholic, smoke, or having a history of (CVD) cardiovascular disease, thyroid disease, and hormonal abnormalities problems were exempted from this study.In the present study SIRT1, TNF-alpha, iNOS were measured by enzyme linked immune absorbent assay (ELISA). Biochemical parameters fasting glucose, body mass index (BMI), urea, creatinine and lipid profile that were measured by spectrophotometer technique in patients and healthy volunteers groups.The Statistical Analysis System- SAS (2012) program was used to effect of difference factors in study parameters. Least significant difference –LSD test (ANOVA) was used to significant compare between means. Estimate of correlation coefficient between difference parameters in this study.The results showed that a highly significant increase in levels of FBG, Urea, creatinine and lipid profile except HDL was decrease levels, BMI was decrease in newly diagnosis DMT1 with healthy volunteers groups. A significant decrease in SIRT1 protein, and a highly significant increase in TNF-alpha and iNOS in newly diagnosis DMT1 with healthy volunteers groups. A highly negative significantly correlation coefficient between SIRT1 and F.G., TG., VLDL,TG/HDL, A.I. and negative significantly correlation duration, urea, creatinine,T.CH., LDL. And non-significant correlation with age, BMI, HDL, CH/HDL and LDL/HDL.. Effect of gender were insignificant of SIRT1. Effect of family history in patients. SIRT1 were all insignificant in patients with family history as with non-family history group. SIRT1 could decreases in the sera of DMT1 patients with family history, the decreased SIRT1 level reflect a mutation in SIRT1 gene, the decreases in SIRT1 on increases the incident of DMT1 complication.

Etiology and prognosis of acute viral encephalitis and meningitis in Chinese children: a multicentre prospective study

BackgroundIn China, there were few studies about the pathogens of acute viral encephalitis and meningitis in children in recent years. The aims of this study were to characterize the etiology and prognosis of acute viral encephalitis and meningitis in Chinese children.MethodsThis was a multicentre prospective study. Two hundred and sixty one viral encephalitis patients and 285 viral meningitis patients were enrolled. The mean age of viral encephalitis and meningitis were 5.88 ± 3.60 years and 6.39 ± 3.57 years, respectively. Real-time reverse transcription PCR and multiplex PCR were used to detect human enteroviruses and herpes viruses in cerebrospinal fluid (CSF) of patients with encephalitis or meningitis. The enzyme-linked immune absorbent assay (ELISA) was used for detecting IgM antibody against Japanese encephalitis virus (JEV) in CSF and against mumps virus, tick-borne encephalitis virus (TBEV), dengue virus and rubella virus in acute serum. The clinical and outcome data were collected during patients’ hospitalization.ResultsThe etiology of viral encephalitis was confirmed in 52.5% patients. The primary pathogen was human enteroviruses (27.7%) in viral encephalitis. The incidence of sequelae and the fatality rate of viral encephalitis with confirmed etiology were 7.5% and 0.8%, respectively. The etiology of viral meningitis was identified in 42.8% cases. The leading pathogen was also human enteroviruses (37.7%) in viral meningitis. The prognosis of viral meningitis was favorable with only 0.7% patients had neurological sequelae.ConclusionsHuman enteroviruses were the leading cause both in acute viral encephalitis and viral meningitis in children. The incidence of sequelae and fatality rate of viral encephalitis with confirmed etiology were 7.5% and 0.8%, respectively. The prognosis of viral meningitis was favorable compared to viral encephalitis.

Should we Investigate Gastroenterology Patients for Pancreatic Exocrine Insufficiency? A Dual Centre UK Study.

Pancreatic exocrine insufficiency may be under recognised in gastroenterological practice. We aimed to identify the prevalence of pancreatic insufficiency in secondary care gastroenterology clinics and determine if co-morbidity or presenting symptoms could predict diagnosis. A secondary aim was to assess response to treatment. A dual centre retrospective analysis was conducted in secondary care gastroenterology clinics. Patients tested for pancreatic exocrine insufficiency with faecal elastase-1 (FEL-1) between 2009 and 2013 were identified in two centres. Demographics, indication and co-morbidities were recorded in addition to dose and response to pancreatic enzyme replacement therapy. Binary logistic regression was used to assess if symptoms or co-morbidities could predict pancreatic insufficiency. 1821 patients were tested, 13.1% had low FEL-1 (<200µg/g). This prevalence was sub-analysed with 5.4% having FEL-1 100-200µg/g (mild insufficiency) and 7.6% having faecal elastase readings <100µg/g. Low FEL-1 was most significantly associated with weight loss or steatorrhoea. Co-morbidity analysis showed that low levels were significantly associated with excess alcohol intake, diabetes mellitus or human immunodeficiency virus; 80.0% treated with enzyme supplements reported symptomatic benefit with no difference in response between high and low dose supplementation (p=0.761). Targeting the use of FEL-1 in individuals with specific symptoms and associated conditions can lead to improved recognition of pancreatic exocrine insufficiency in a significant proportion of secondary care patients. Intervening with lifestyle advice such as smoking cessation and minimising alcohol intake could improve outcomes. In addition, up to 80% of patients with low faecal elastase respond to supplementation.

Screening of aflatoxin M1 occurrence in selected milk and dairy products in Terengganu, Malaysia

The study was conducted to screen the occurrence of aflatoxin M1 (AFM1) in 53 selected milk and dairy product samples (11 liquid milk, 12 powdered milk, 8 3-in-1 beverages, 6 condensed sweetened milk, 2 evaporated milk, 7 cultured milk drink, 5 yogurt and 2 cheese samples). These samples were purchased from selected markets in Terengganu, Malaysia in January 2014 based on a questionnaire survey among 212 respondents on the types and brands of milk and dairy products that were frequently consumed. Based on the responses, 53 milk and dairy products were purchased and the competitive enzyme-linked immune-absorbent assay (ELISA) method was used to determine the level of AFM1 in the samples. Of 53 samples, 19 samples were positive with AFM1 (35.8%) ranging from 3.5 to 100.5 ng/L. Although 4/53 (7.5%) of the tested samples had the contamination level greater than the European Commission (EC) limit (>50 ng/L), the contamination levels were still below the Malaysia Food Regulation 1985 limit (less than 500 ng/L). This study provided a pioneering data on the occurrence of AFM1 in milk and dairy products in Malaysia.

Delivery of Topically Applied Calpain Inhibitory Peptide to the Posterior Segment of the Rat Eye.

We developed an inhibitory peptide that specifically acts against mitochondrial μ-calpain (Tat-μCL, 23 amino acid, 2857.37 Da) and protects photoreceptors in retinal dystrophic rats. In the present study, we topically administered Tat-μCL to the eyes of Sprague-Dawley rats for 7 days to determine both the delivery route of the peptide to the posterior segment of the eye and the kinetics after topical application in adult rats. Distribution of the peptide was determined by immunohistochemical analysis, and enzyme-linked immune-absorbent assay was used to quantify the accumulation in the retina. Peptides were prominently detected in both the anterior and posterior segments of the eye at 1 h after the final eye drop application. Immunohistochemically positive reactions were observed in the retina, optic nerve, choroid, sclera and the retrobulbar tissues, even in the posterior portion of the eye. Immunoactivities gradually diminished at 3 and 6 h after the final eye drop. Quantitative estimations of the amount of peptide in the retina were 15.3, 5.8 and 1.0 pg/μg protein at 1, 3 and 6 h after the final instillation, respectively. Current results suggest that while the topically applied Tat-μCL peptide reaches the posterior segment of the retina and the optic nerve, the sufficient concentration (> IC50) is maintained for at least 6 h in the rat retina. Our findings suggest that delivery of topically applied peptide to the posterior segment and optic nerve occurs through the conjunctiva, periocular connective tissue, sclera and optic nerve sheath.

Serum thrombopoietin levels and its relationship with thrombocytopenia in patients with cirrhosis.

Background:Patients with cirrhosis usually have thrombocytopenia in discrete levels. The mechanism of thrombocytopenia is thought as splenic sequestration and destruction of platelets, impaired bone marrow generation and diminished hepatic thrombopoietin synthesis.Objectives:The aim of this study was to evaluate serum thrombopoietin levels and its relationship with thrombocytopenia at patients with cirrhosis.Patients and Methods:Ninety–two cirrhotic patients and 45 healthy controls without history or findings of pathologies that can effect thrombopoietin levels were enrolled by simple random sampling to patient and control groups of this case control study performed at Eskisehir-Turkey. Thrombopoietin was measured in serum samples with a solid phase enzyme-linked immune absorbent assay. Additionally, spleen size and volume index were determined.Results:Platelet counts were lower in patients with cirrhosis (97000 ± 8000/mm3) than in healthy subjects (240000 ± 51000/mm3, P < 0.001). Significant difference was determined for platelet counts among child A, B and C stages (Child A vs. Child B P < 0.05 Child A vs. Child C P < 0.001–Child B vs. Child C P < 0.05). Serum TPO concentration was higher (69 ± 12 pg/mL) in cirrhotic group than healthy controls (49 ± 9 pg/ml) (P < 0.05). No significant difference in TPO levels were found among the Child A, B and C stages (64 ± 11 pg/mL, 75 ± 13 pg/mL and 68 ± 10 pg/mL, respectively). Spleen size and SVI was significantly higher in the cirrhotic patients than healthy controls (148 ± 14 mm vs. 98 ± 11 mm, P < 0.001-9167 ± 287 cm2 vs. 4118 ± 123 cm2). Significant difference was determined for spleen size and spleen index among child A, B and C stages (Child A vs. Child B P < 0.05 Child A vs. Child C P < 0.001–Child B vs. Child C P < 0.05). TPO levels were significantly different between cirrhotic patients with platelet levels below 50.000/mm3 (n = 16, plt-count: 41000 ± 8300/mm3, TPO levels: 73 ± 7 pg/mL) and above 50.000/mm3 (n = 76, plt-count: 105000 ± 9500/mm3, TPO levels: 65 ± 10 pg/mL) (P < 0.01). In correlation analysis, there was a strong negative correlation between platelet count-spleen size (P < 0.001, r = -0.74) and platelet count–serum TPO levels (P < 0.001, r = -0.71).Conclusions:Our results suggest that liver cirrhosis does not cause impaired thrombopoietin production even in the late stage of disease. Thrombopoietin has no contribution for the occurrence of thrombocytopenia in cirrhosis; splenic sequestration seems to be the main factor.

Humoural immune response and pathological analysis in patients with false immune diagnosis of cystic echinococcosis

The patients with false immune diagnosis of hydatid disease were investigated for the humoural immune response to analyse the possible reasons and mechanism leading to false immune diagnosis. Two hundred and thirty-nine patients with nature-unknown cysts and 30 healthy controls were detected by immunological assays (four hydatid antigen-based immunogold filtration assay and enzyme-linked immune absorbent assay) and ultrasound. Sensitivity of and specificity of immunological assay and ultrasound were calculated, respectively. The serological diagnosis was compared with surgical pathology to screen the patients with false immune diagnosis for the immunoglobulin measurement and pathological analysis. The history and cyst characteristics were also reviewed. The results indicate the immunoglobulin has little influence on false immunodiagnosis. The false-negative immunodiagnosis was caused by the cysts' inactive status while the false positive caused by previous rupture, antigen cross-reaction. The clinical diagnosis of cystic echinococcosis requires a combination of immunodiagnosis and ultrasonography, which is the necessary complementary confirmation.

Prostaglandin E2 alteration in contraceptive consumers: as a risk factor for inflammatory diseases

The aim of this study was to evaluate the effect of oral combined contraceptive pills on Prostaglandin E2 levels and lipid profiles. The enzyme-linked immune absorbent assay method and spectrophotometric assay were used for the evaluation of PGE2 levels and lipid profiles, respectively, in 50 healthy women with normal menstrual cycles who served as the control group and 50 women taking contraceptive pill. The data obtained for serum Prostaglandin E2, LDL-C, and cholesterol concentrations in contraceptive pill consumers were significantly upper (P=0.04, 0.002, and 0.05, respectively) than control group. The age of contraceptive pill consumption and the duration of pill intake beyond 36months had no significant effect on the prostaglandin E2 concentration. It is suggested that the increase of Prostaglandin E2 and atherogenic lipid levels may be related to their probable effects in response to various pathological and physiological properties of COCs.

Elevated serum and tissue VEGF associated with poor outcome in breast cancer patients

Vascular endothelial growth factor (VEGF) has a potent angiogenesis functions in experimental models, although their role in the progression of human breast cancer is unclear. The aim of the current study was to examine the expression pattern of VEGF in serum and tissues of breast cancer patients, examine the tumor vascular characteristic by counting the blood vessels to assess microvessles density (MVD) and conduct correlations between the expressions of growth factor in relation to patient's clinicopathological data and survival. MethodsOne hundred and twenty untreated patients with breast cancer were included in the study and followed for 4 years and 30 females with benign breast lesions matched with age and menstrual state as (control group). In this work we examine serum and tissue expression of VEGF by enzyme linked immune absorbent assay (ELISA) and immunoperoxidase technique respectively. Microvessels density were assessed and correlated with expression of growth factors. ResultsThe mean serum level of VEGF elevated in breast cancer patients before surgery was significantly higher when compared to that in patients with benign breast lesions or in the same patient after surgery. There was positive correlation between serum and tissue VEGF. Serum and tissue vascular endothelial growth factor was strongly associated with grade III tumor, large tumor size, positive lymph node, negative hormone receptor status, +ve HER2 neu and poor survival, the data of the present study showed significant increase in mean serum level of VEGF in patients with positive vascular invasion P: 0.013. ConclusionVEGF appear to play an important role in progression of breast carcinoma and to have significant impact on patient prognosis and can be used to identify a subset of breast cancer at higher risk for development of recurrence and distant metastasis.