Lactate Deshydrogenase Research Articles

Assessment of the antidiabetic and antilipidemic properties of Bacillus subtilis SPB1 biosurfactant in alloxan-induced diabetic rats.

The present study aimed to scrutinize the potential of Bacillus subtilis SPB1biosurfactant, orally administered, for preventing diabetic complications in rats. The findings revealed that, Bacillus subtilis biosurfactant was an effective reducer of α-amylase activity in the plasma. Moreover, this supplement helped protect the β-cells from death and damage. Both the inhibitory action of SPB1 biosurfactant on α-amylase and the protection of the pancreas' β-cells lead to a decrease of the blood glucose levels, consequently antihyperglycemic effect. Interestingly, this lipopeptide biosurfactant modulated key enzyme related to hyperlipidemia as lipase; which leads to the regulation of the lipid profile in serum by the delay in the absorption of LDL-cholesterol and triglycerides, and a significant increase in HDL-cholesterol. Histological analyses also showed that it exerted a protective action on the pancreases and efficiently preserved the liver-kidney functions of diabetic rats, evidenced by significant decreases in aspartate transaminase, alanine transaminase, gamma-glytamyl transpeptidase and lactate deshydrogenase activities in the plasma, as well as in the creatinine and urea contents. Overall, the present study demonstrated that the hypoglycemic and antilipidemic activities exhibited by Bacillus subtilis biosurfactant were effective enough to alleviate induced diabetes in experimental rats. Therefore, SPB1biosurfactant could be considered as a potential strong candidate for the treatment and prevention of diabetes.

TSPO is a new anti-inflammatory target in the airway of COPD

Introduction Chronic obstructive pulmonary disease (COPD) is a major public health problem. It is well-known that COPD is characterized by a systemic inflammatory response. Recent studies have shown that the 18 kDa translocator protein (TSPO) – also known as the peripheral benzodiazepine receptor, plays a key role in the regulation of immune function and inflammation. Benzodiazepine use is high among adults with COPD. The aim of this study was to investigate TSPO expression in COPD biopsies and to establish an in vitro model to examine the role of TSPO in COPD inflammation. Materials and methods Samples were obtained from lung surgery of healthy-smokers and COPD patients. TSPO expression was evaluated using immunohistochemistry and morphometric analysis. TSPO agonist (Ro5-4864) and antagonist (PK-11195) were used on NCI-H292 and air liquid interface (ALI) cultures of bronchial epithelial cells. Cellular TSPO expression was assessed by PCR. Cigarette smoke extract (CSE)-induced epithelial inflammation and cell viability were assessed by interleukin 8 (IL-8) and lactate deshydrogenase assays respectively. Results TSPO expression was mainly detected in bronchial epithelium and inflammatory cells. Expression levels were independently associated with COPD status (P-value 0.01827) and age (P-value 0.00203). There was no association with benzodiazepine intake. NCI-H292 and airway epithelial cells cultivated at ALI both expressed TSPO. In NCI-H292, Ro5-4864 (10−9 M) increased IL-8 release. PK-11195 (10−9 M) partially prevented CSE-induced IL-8 release. Conclusion TSPO appears to play an important role in COPD airway epithelial cells. TSPO antagonist PK-11195 decreased CSE-induced inflammation. TSPO antagonist should be further evaluated as a potential future COPD treatment.

A New Prognostic Score for Patients Older Than 60 Years with a Classical Hodgkin Lymphoma: A Retrospective Analysis on 165 Patients

Abstract Background: Patients (pts) with classical Hodgkin Lymphoma (CHL) older than 60 years represent about 15% to 20% of CHL pts in population registries. Elderly CHL treated with standard protocols always have a poorer outcome as compared to younger pts. The aim of this retrospective study is to identify specific prognostic factors in a cohort of 165 CHL pts aged over 60 years. Methods: From two hospital registries in Lyon (France), we analyzed 165 CHL pts older than 60 years and treated between 1981 and 2012. None were HIV positive. The following clinical characteristics were recorded: age, gender, presence of B-symptoms, ECOG performance status (PS), Cumulative Illness Rating Scale (CIRS), Ann Arbor stage, bone marrow involvement, complete blood counts, lactate deshydrogenase(LDH), C-reactive protein (CRP), fibrinogen, serum albumin level and protein electrophoresis. International Prognostic Score (IPS) was collected for 116 pts (70%). A total of 136 pts (82%) were treated with an anthracycline based chemotherapy (CT) and 53 pts (32%) received radiation therapy. Clinical and biological characteristics at diagnosis were assessed by a Cox regression for univariate and multivariate analysis for progression free survival (PFS) and overall survival (OS). We then designed a prognostic score for PFS and OS based on significant variables from multivariate analysis at threshold p-value < 0.5 and compared the effectiveness of these scores to IPS by a receiver operating characteristic (ROC) analysis. Results: The median age at diagnosis was 69 years (range, 60-89). 60% were male, 65% presented with B-symptoms, 38% had a PS> 2 and 16% had at least one comorbidity scored 3 or 4 on CIRS. 56% had an Ann Arbor stage III-IV.65% of pts achieved a complete or a partial response, 16% were refractory or progressed within 6 months after the start of treatment and 19% died during treatment. Median follow up is 65 months (1.1-207.6). 47 pts (29%) have progressed or relapsed. Median duration of PFS is 39.6 months (95%CI: 23.9-73.3) with a 4-year PFS rate of 49% (95%CI: 40-56). 78 pts (47%) died, 22% within the first year after treatment initiation. Median duration of OS is 73.3 months (95%CI: 38.5-129.8) and the 4-year OS rate is 55% (95%CI: 47-63). The main causes of death were HL progression (39%), infection (17%), treatment toxicity (13%) and second cancer (10%). Pts treated with or without an anthracycline based CT did not have significantly different 4-year PFS (50% vs. 43%, P=0.55) or OS (57% vs. 47%, P=0.22). In univariate analysis, age, B-symptoms, PS, CIRS, albumin, hemoglobin, lymphocyte count, bone marrow involvement, Ann Arbor stage, protein, gammaglobulin, fibrinogen, LDH and CRP were associated with a shortened PFS. Factors associated with a shortened OS were: sex, age, PS , CIRS, albumin, hemoglobin, lymphocyte count, bone marrow involvement, Ann Arbor stage, protein, CRP, gammaglobulin and fibrinogen. In multivariate analysis, age>69 years (HR=3.97; 95%CI: 2.11-7.47, P<0.0001), bone marrow involvement (HR=2.09; 95%CI: 1.11-3.91, P=0.02), gammaglobulin level<10g/L (HR=2.43; 95%CI: 1.31-4.50, P=0.005) and fibrinogen level>5g/L (HR=2.29; 95%CI: 1.25-4.19, P=0.007) were independently associated with a shortened PFS. Pts with 0-1, 2 and 3-4 adverse factors had a median duration of PFS of 180, 37 and 4.6 months respectively (Figure, P=<0.0001). In multivariate analysis, factors independently associated with a poor OS were age>69 years (HR=3.66; 95%CI: 1.95-6.85, P<0.0001), PS > 2 (HR=3.04; 95%CI: 1.69-5.45, P=0.0002) and gammaglobulin level<10g/L (HR=2.26; 95%CI: 1.25-4.07, P=0.007). Pts with 0, 1, 2 and 3 adverse factors had a median duration of OS of 180, 73, 21.5 and 3 months respectively (P<0.0001). Eventually, evaluation of our model by ROC analysis showed a better predictive value than IPS for PFS (0.69 vs. 0.54, P=0.01) and OS (0.74 vs. 0.60, P=0.02) . Conclusion: This prognostic score identifies subgroups of elderly CHL pts at high risk of progression and early death, using simple biological and clinical parameters at diagnosis. Bone marrow involvement is one of the factors independently associated with PFS. This result supports bone marrow evaluation at diagnosis. This score better discriminates elderly CHL pts’ outcome compared to IPS. Our results should be confirmed in independent and prospective studies in combination with complete geriatric assessment. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Efficacy of Orally VP16 Plus Prednisone with or without Cyclophosphamide for Refractory and Relapsed Peripheral T-Cell Lymphoma: A Retrospective Single Center Experience

Background: Peripheral T-cell lymphoma represents a heterogeneous group of mature T/NK lymphoma with an aggressive presentation course and a poor outcome. Disease history is characterized by a high degree of relapsed or refractory disease and an inferior outcome. Etoposide phosphate (VP16) is a non-specific antineoplastic agent frequently used in combination in oncology since 50 years. The major mechanism of action of etoposide is the inhibition of DNA topoisomerase II involved in the unwinding of the DNA molecule during replication. A recent preclinical study suggests an immunomodulatory effect of etoposide by selectively eliminating activated T cells. Specific efficacy of VP16 in T-cell lymphoma and especially in AITL had not been described.Methods: We retrospectively analyzed data on adult patients with refractory or relapsed angioimmunoblastic T-cell lymphoma (AITL), anaplastic large cell lymphoma and peripheral T-cell lymphoma not otherwise specified (PTCL-NOS) treated in the university hospital center of Tours (France) by VP16 with or without cyclophosphamide between January 1992 and June 2014. VP16 was administrated orally at 75mg daily continuously or at 100 mg/m² daily for 3 days in 15-day cycles if it was associated with oral cyclophosphamide (150mg/m² daily for 3 days). Treatment was associated with prednisone 60 mg/m²/day. All responses were described according to Cheson criteria (2007). Adverse event were reported according to CTCAE (Common Toxicity Criteria of Adverse Events). Treatment was prescribed until progressive disease, intensive therapy or death, and could be stopped according to clinician appreciation after prolonged remission. Overall survival (OS) and event-free survival (EFS) were estimated since time to initiation of VP16 using the method of Kaplan and Meier. Event was defined as disease progression, death or change of treatment for any cause (toxicity, allograft etc.).Results: A total of 23 patients were enrolled : 13 AITCL, 2 anaplastic lymphoma (all of them were ALK-negative) and 8 PTCL-NOS. Median age at diagnostic was 57.3 years (range, 31-80) with a male preponderance (14/9). Most patients had B-symptoms (n=14) and advanced stage (n=19) with more than one extranodal site for 10 patients. Lactate deshydrogenase was upper normal value for 14 patients. The median number of prior systemic therapies was two (range, 2 to 4). Five patients received VP16 with cyclophosphamide. The overall response rate (ORR) was 65.2% (15 of 23), including 39.1% complete response (CR) (9 of 23). With a median follow up of 64.8 months (range, 33.7 to 96), the median OS was 39 months (95% IC: 4 months-not reached), and EFS 5 months (95% IC: 3 to 39) (Figure 1 and 2). Median duration of treatment was 15 months (range, 1 to 91). For AITL, the ORR was 69.2% (9 of 13), including 46.2% CR (6 of 13). The median OS was 77 months (95% IC : 5 months-not reached), and EFS 39 months (95% IC : 3-73). One haematological adverse event ≥grade 3 induced end of treatment, none extra-haematological adverse event was reported. Three patients received intensive regimen follow by allogenic stem cells transplantation and two of them (both AITL) after the response obtained with VP16. Progressive disease was the main cause of end of treatment (n=15) and of death (n=9). Patients treated with VP16 and cyclophosphamide had the same outcome than those treated with VP16 alone. For the 9 patients who reached a CR, 7 are still under CR at the end of follow up with median time of response of 67.4 months (range, 3 to 244).Conclusion: With or without cyclophosphamide, VP16 seems to be an effective and safe treatment for relapsed and refractory PTCL. AITL could have a higher sensibility to VP16 than other subtypes with durable responses. A prospective study should confirm these results, even for patient eligible for transplant program. Finally, orally VP16 can be an effective alternative treatment for unfit patients who cannot be included in clinical studies evaluating novel agents for PTCL. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Nitrate-induced Biochemical and Histopathological Changes in the Liver of Rats: Ameliorative Effect of Hyparrhenia hirta

ObjectiveThe present study investigated the protective role of Hyparrhenia hirta (H. hirta) against sodium nitrate (NaNO3)-induced hepatoxicity. MethodsMale Wistar rats were randomly divided into three groups: a control group and two treated groups during 50 d with NaNO3 administered either alone in drinking water or co-administered with H. hirta. ResultsNaNO3 treatment induced a significant increase in serum levels of glucose, total cholesterol and triglyceride while serum total protein level decreased significantly. Transaminases and lactate deshydrogenase activities in serum were elevated indicating hepatic cells' damage after treatment with NaNO3. The hyperbilirubinemia and the increased serum gamma glutamyl transferase activities suggested the presence of cholestasis in NaNO3 exposed rats. In parallel, a significant increase in malondialdehyde level along with a concomitant decrease in total glutathione content and superoxide dismutase, catalase and glutathione peroxidase activities were observed in the liver after NaNO3 treatment. Furthermore, nitrate caused a significant induction of DNA fragmentation. These modifications in NaNO3-treated rats corresponded histologically with hepatocellular necrosis and mononuclear cells infiltration. H. hirta supplementation showed a remarkable amelioration of the abnormalities cited above. ConclusionThe results concluded that the treatment with H. hirta had a significant role in protecting the animals from nitrate-induced liver dysfunction.

Changes in Oxidative Stress Markers and Biological Markers of Muscle Injury with Aging at Rest and in Response to an Exhaustive Exercise

PurposeThe aim of this study was to evaluate whether oxidative stress markers and biomarkers of muscle injury would be affected by aging at rest and in response to an incremental exhaustive exercise.MethodsFifteen young (20.3±2.8 years) and fifteen older adults (65.1±3.5 years) performed an incremental cycle ergometer test to exhaustion. Before and after exercise, oxidative stress [superoxide dismutase (SOD), glutathione peroxidase (GPX), glutathione reductase (GR), ascorbic acid, α-Tocopherol, malondialdehyde (MDA)] and muscle injury [creatine kinase (CK), lactate deshydrogenase (LDH)] biomarkers were assessed.ResultsAt rest, there was no difference in oxidative stress markers and LDH level between the groups, however CK was significantly higher in the young group than the elderly group (p<0.05). During recovery, in comparison with resting values, a significant increase in SOD (1092±145.9 vs. 1243±98 U/g Hb), GPX (67.4±12.7 vs. 79.2±15.6 U/g Hb) and GR (6.5±0.9 vs. 7.7±0.5 U/g Hb) activities were observed only in the young group (p<0.05). MDA has increased only in the older group (0.54±0.2 vs. 0.79±0.2 µmol/l) (p<0.01). CK increased in both groups (young group: 122.5±22.2 vs. 161.9±18.7 UI/l; older group: 88.8±34.1 vs. 111.1±25.9 UI/l) (p<0.01), however LDH has increased only in the young group (400.5±22.2 vs. 485±18.7 UI/l) (p<0.01) without alteration in the older group (382.8±34.1 vs. 418.5±25.9 UI/l).ConclusionsThese findings indicate that aging is associated with a decrease in antioxidant efficiency and an increase in oxidative stress damage. Furthermore, older adults would not more susceptible to exercise-induced muscle injury than young people.

Évaluation de l’effet cardioprotecteur de l’extrait aqueux de Mareya micrantha (Euphorbiaceae) chez le lapin

We investigated the cardioprotective effect of Mareya micrantha (Euphorbiaceae) from the dosage of 3 biochemical markers. It is a plant traditionally used as laxative, antibacterial and for abortion affections in Côte d’Ivoire and in other countries in West Africa. Increasing doses of aqueous crude extract of Mareya micrantha were injected to various rabbits of different batches. Then we evaluated the variation of the serum activities of glutamo-oxaloacetic transaminase (GOT), lactate-deshydrogenase (LDH) and creatinephosphokinase (CPK). The statistic analysis of results shows a reduction of serum activities of CPK and LDH (P < 0.05) but it indicates an insignificant variation of the serum activity of GOT (P > 0.05). In conclusion, we can say the extract of Mareya micrantha with the dose of 100 mg/kg of body weight during 4 weeks is well tolerated by heart and it also may have heart protector effect in rabbits.

Picea mariana polyphenolic extract inhibits phlogogenic mediators produced by TNF-α-activated psoriatic keratinocytes: Impact on NF-κB pathway

Ethnopharmacological relevancePicea mariana ((Miller) Britton, Sterns, and Poggenburg; Pinaceae) bark has been traditionally used by North American natives for treating topical inflammations. It has been also suggested to improve various inflammatory skin disorders like Psoriasis vulgaris. Extracts from this bark storage protein contain polyphenolic compounds which have well-known antiinflammatory activities. Based on the capacity of polyphenolic compounds to modulate functions of normal human keratinocytes, this study was set up to decipher the mechanisms of action of a chemically characterized polyphenolic extract from Picea mariana bark (BS-EAcf) on lesional keratinocytes of skin with psoriasis vulgaris, a disease driven by the immune system in which TNF-α plays a significant role. Materials and methodsBS-EAcf corresponds to the ethyl acetate soluble fraction from the hot water extract of Picea mariana bark. BS-EAcf effects were evaluated in normal human (NHK) and psoriatic (PK) keratinocytes stimulated by TNF-α. Cell viability was assessed by lactate deshydrogenase release and propidium iodide (PI) staining. The mechanisms of action of BS-EAcf in keratinocytes were investigated by flow cytometry, ELISAs, RT-PCR and western blot analyses. ResultsPK exhibited a higher response to TNF-α than NHK regarding the ICAM-1 expression and the production of NO, IL-6, IL-8, fractalkine and PGE2, whereas BS-EAcf significantly inhibited this TNF-α-induced increase at concentrations without causing keratinocyte toxicity. Additionally, this extract significantly inhibited the TNF-α-induced release of elafin and VEGF by PK and NHK. Since TNF-α activation of most of these factors is dependent on the NF-κB pathway, this latter was studied in TNF-α-activated PK. BS-EAcf inhibited the TNF-α-induced phosphorylation and degradation of total IκBα as well as phosphorylation of NF-κB p65. ConclusionsThe ethyl acetate fraction from Picea mariana bark extract showed inhibitory effects of cytokines, chemokines, adhesion molecules, nitric oxide and prostaglandins produced by keratinocytes under TNF-α activation through down-regulating the NF-κB pathway. This study demontrated that this extract could be a potential antiinflammatory agent capable of improving psoriatic skin.

Perirenal fibrosis: make your diagnosis.

A 72-year-old patient treated with BCG immunotherapy for relapse of urothelial cancer was admitted for diffuse oedema. His past history was characterized by recurrent pleuropericarditis treated by pericardotomy 16 years earlier and by retroperitoneal fibrosis (RPF) treated by steroids 15 years earlier. Serum creatinine rose progressively from 190 to 618 μmol/L and he was admitted in the nephrology department. Clinical examination showed diffuse oedema, dyspnoea and mild tibial tenderness. Haemoglobin was 99 g/L with normal platelet and leukocyte counts. Calcaemia, phosphoraemia, magnesaemia, haptoglobin and lactate deshydrogenase levels were within the normal range. Serum was negative for antinuclear and antineutrophil cytoplasm antibodies. Bilateral pyelic dilatation was diagnosed and treated by bilateral ureteral stenting. Echocardiography, cardiac magnetic resonance imaging (MRI) and right cardiac catheterization confirmed restrictive cardiopathy related to constrictive pericarditis with severe thickening of the lateral wall of the pericardium (not shown). An abdominal CT scan was performed and is shown in Figure 1.

Methylene Blue Protects Liver Oxidative Capacity after Gut Ischaemia–Reperfusion in the Rat

ObjectivesMesenteric ischaemia/reperfusion (IR) may lead to liver mitochondrial dysfunction and multiple organ failure. We determined whether gut IR induces early impairment of liver mitochondrial oxidative activity and whether methylene blue (MB) might afford protection. DesignControlled animal study. Materials and methodsRats were randomised into three groups: controls (n = 18), gut IR group (mesenteric ischaemia (60 min)/reperfusion (60 min)) (n = 18) and gut IR + MB group (15 mg kg−1 MB intra-peritoneally) (n = 16). Study parameters were: serum liver function markers, blood lactate, standard histology and DNA fragmentation (apoptosis) on intestinal and liver tissue, maximal oxidative capacity of liver mitochondria (state 3) and activity of complexes II, III and IV of the respiratory chain measured using a Clark oxygen electrode. ResultsGut IR increased lactate deshydrogenase (+982%), aspartate and alanine aminotransferases (+43% and +74%, respectively) and lactate levels (+271%). It induced segmental loss of intestinal villi and cryptic apoptosis. It reduced liver state 3 respiration by 30% from 50.1 ± 3 to 35.2 ± 3.5 μM O2 min−1 g−1 (P < 0.01) and the activity of complexes II, III and IV of the mitochondrial respiratory chain. Early impairment of liver mitochondrial respiration was related to blood lactate levels (r2 = 0.45). MB restored liver mitochondrial function. ConclusionsMB protected against gut IR-induced liver mitochondria dysfunction.

Children with Sickle Cell Anemia Experience Severe Oxygen Desaturation During Night and After Six-Minute Walk Distance Test

Abstract 4766 Background:Respiratory complications are the first causes of death among adult patients with sickle cell anemia (SCA). Finding risk factors for children is important. We study clinical, biological, respiratory and heart parameters, as well as exercise and sleep oxygen saturation in SCA children. Patients and Methods:We conducted a prospective study in homozygous SS or S/beta 0 thalassemic children. A chronic transfusion program was an exclusion criterion. We recorded the number of vaso-occlusive crises (VOC) the year before and after inclusion, past history of acute chest syndrome (ACS), hydroxyurea treatment, tonsils size, baseline heart rate and blood pressure, baseline hemoglobin (Hb), reticulocytes, fetal Hb levels, leukocytes and platelets counts, total bilirubin, aspartate aminotransferase (AST), lactate deshydrogenase (LDH). All patients underwent respiratory function testing (RFT), echocardiography assessing tricuspid regurgitation velocity jet (TRV). We measured daytime oxygen saturation using a Radical Masimo set ® pulse oximeter. All patients underwent a non-encouraged six-minute walk test (6MWT). Nocturnal pulse oximetry was recorded using a Nonin ® device during 3 consecutive nights for 30 patients. We considered the average night-time oxygen saturation and the percentage of sleep time with oxygen saturation less than 90%. Statistical analysis was conducted using the Fisher exact test for categorical variables and the Wilcoxon test for continuous variables. Results:Forty-two unselected SCA children were enrolled. Three patients were secondarily excluded because the echocardiography revealed asymptomatic cardiac anomalies (two pulmonary valve stenosis and one persistent arterial canal). In the remaining 39 patients, 38 were SS and one was S/beta 0 thalassemic. Median age was 10.8 years (range 5.7–17); 25 patients were females (64%). The median number of VOC was 0 the year before inclusion (range 0–6), and 0 the year after (range 0–7). Fifteen patients (38%) had displayed at least one ACS. Nine patients (23%) were receiving hydroxyurea treatment. Sixteen patients (43%) had tonsillitis enlargement. Median basal heart rate was 97 bpm (range 75–122). Mean systolic blood pressure was 107 ± 11.3 mm Hg and mean diastolic blood pressure was 64 ± 6.6 mm Hg. Mean Hb was 7.9 ± 1.2 g/dL, mean reticulocyte count was 236 ± 82 Giga/L, median HbF was 9.2 % (range 0.8–28), mean leukocyte count was 11.1 ± 3.2 Giga/L, mean platelet count was 407 ± 132 Giga/L. Median total bilirubin, AST, and LDH were, respectively, 41.5 mg/dL (range 13–163), 62 UI/l (range 35–132), and 1421 UI/l (range 618–1893) (normal range for LDH in our lab 125–243). Fifteen patients (38.5%) had abnormal RFT: 4 had obstructive pattern, 3 had restrictive pattern, and 3 had both. Left ventricular diastolic function was normal for all patients. Six patients had a TRV above 2.6 m/s. Median daytime oxygen saturation was 97 % (range 89–100). One patient had a daytime saturation below 92%. Median nocturnal oxygen saturation was 94.7 % (range 87.7–99.5). Ten patients (33%) displayed average night-time saturation below 92%. Eleven patients (37%) spent more than 10% of their sleep time with oxygen saturation below 90%. Mean six-minute walk distance (6MWD) was 547 ± 99 m. After the 6MWT, 14 patients (35%) had an oxygen saturation below 92%. Median difference in oxygen saturation before and after the test was 2% (range −57, +2). Nocturnal hypoxemia was not associated with age, gender, tonsils size, hydroxyurea treatment, past history of ACS, RFT pattern, number of VOC, leukocytes, platelets, LDH, bilirubin nor AST. It was associated with Hb level (7.2±1.2 g/dL if nocturnal hypoxemia vs 8.4±1.1, p=0.02), daytime oxygen saturation (94% [range 92–99] if nocturnal hypoxemia vs 98% [range 89–100], p=0.03), and oxygen saturation after 6MWT (91% [range 40–99] if nocturnal hypoxemia vs 96% [range 79–100], p=0.03). Children with a TRV above 2.6m/s had a significantly lower Hb level (7.4 g/dL [6.4–8.1] vs 8.5 [6.5–10.6]). Conclusions:Our study emphasizes the frequency of night-time oxygen desaturation in SCA children. It shows that a simple effort can induce a significant decrease in oxygen saturation. The consequences of hypoxemia are difficult to assess given the small sample size. One can hypothesize that hypoxemia and hypoxia/reoxygenation cycles both contribute to the pathophysiology of the disease through inflammation and vascular injury. Disclosures:No relevant conflicts of interest to declare.

Attenuation of endoplasmic reticulum stress and mitochondrial injury in kidney with ischemic postconditioning application and trimetazidine treatment

BackgroundEndoplasmic reticulum (ER) and mitochondria have been implicated in the pathology of renal ischemia/reperfusion (I/R). In the present study, we investigated whether the use of ischemic postconditioning (IPostC) and trimetazidine (TMZ) separately or combined could reduce ER stress and mitochondria damage after renal ischemia.MethodsKidneys of Wistar rats were subjected to 60-min of warm ischemia followed by 120-min of reperfusion (I/R group, n = 6), or to 6 cycles of ischemia/reperfusion (10-s each cycle) just after 60-min of warm ischemia (IPostC group, n = 6), or to i.p. injection of TMZ (3 mg/kg) 30-min before ischemia (TMZ group, n = 6), or to the combination of both treatments (IPostC+TMZ group, n = 6). The results of these experimental groups were compared to those of a sham-operated group in which rat renal pedicles were only dissected. Sodium reabsorption rate, creatinine clearance lactate deshydrogenase (LDH) activity in plasma, and concentration of malonedialdehyde (MDA) in tissue were determined. In addition, Western blot analysis was performed to identify the amounts of cytochrome c, c-JunNH2-terminal kinase (JNK), voltage-dependent anion channel (VDAC), glycogen synthase kinase 3-beta (GSK3-β), and ER stress parameters.ResultsIPostC or/and TMZ significantly decreased cytolysis, oxidative stress and improved renal function in comparison to I/R group. IPostC but not TMZ significantly attenuated ER stress parameters versus I/R group. Indeed, it down-regulated the glucose-regulated protein 78 (GRP78), the activating transcription factor 4 (ATF4), the RNA activated protein kinase (PKR)-like ER kinas (PERK), the X box binding protein-1 (XBP-1) and the caspase12 protein levels. TMZ treatment significantly augmented GSK3-β phosphorylation and reduced levels of cytochrome c and VDAC phosphorylation in comparison to IPostC application. The combination of both treatments gave a synergetic effect. It significantly improved the survival rate, attenuated cytolysis, oxidative stress and improved renal function.ConclusionThis study revealed that IPostC protects kidney from I/R injury by suppressing ER stress while the beneficial effects of TMZ are mediated by mitochondria protection. The combination of both treatments ameliorated functional recovery.

Heightened expression of MICA enhances the cytotoxicity of NK cells or CD8+T cells to human corneal epithelium in vitro

BackgroundMajor-histocompatibility-complex class I-related chain A (MICA) antigens are the ligands of NKG2D, which is an activating or coactivating receptor expressed on human NK cells and CD8+T cells. We sought to determine whether MICA expression in human corneal epithelium (HCE) could affect the cytotoxicity mediated by NK cells or CD8+T cells.MethodsCell cultures of HCE were harvested from human donor eyes. Flow cytometric analysis and ELISA was performed to determine the levels of MICA expression on HCE. Then, HCE was transfected with a lentivirus vector expressing MICA and GFP. Flow cytometric analysis, RT-PCR, western blot and ELISA were performed to check the levels of MICA expression. For cytotoxicity testing, allogeneic NK cells and CD8+T cells were isolated from peripheral blood mononuclear cells of healthy volunteers by magnetic cell sorting. The cytolytic activity of NK cells and CD8+T cells was assessed against MICA-transfected HCE (NK cells: E:T ratio = 3:1; CD8+T cells: E:T ratio = 10:1) using the nonradioactive cytotoxicity detection kit lactate deshydrogenase.ResultsSurface expression of MICA on corneal epithelium was identified at a low level. A cell line of stable human MICA-transfected corneal epithelium was successfully established. Heightened expression of MICA on HCE was found to promote the cytotoxicity mediated by NK cells or CD8+T cells, which could be blocked by an anti-MICA antibody.ConclusionMICA molecules may contribute to cytotoxic responses mediated by activated immune effector cells in corneal epithelium immunity.

Cisplatin alone or combined with gemcitabine in carcinomas of unknown primary: Results of the randomised GEFCAPI 02 trial

PurposeTo compare the overall survival rates of good-prognosis carcinomas of an unknown primary site (CUPS) patients treated with cisplatin alone (C) or in combination with gemcitabine (CG). Patients and methodsGood prognosis was defined according to the GEFCAPI (Groupe d’Etude Français des Carcinomes de site Primitif Inconnu) classification by PS (Performance Status) ⩽1 and LDH (Lactate Deshydrogenase) within the normal range. Patients were randomly assigned to receive C or CG. Patients in the C arm received cisplatin 100mg/m2 repeated every 3weeks. In the CG arm, chemotherapy consisted of gemcitabine 1250mg/m2 on days 1 and 8 and cisplatin 100mg/m2 IV on day 1, repeated every 3weeks. The original plan was to accrue 192 patients in order to detect a 20% difference in overall survival. ResultsFifty-two patients were enrolled (arm A: 25; arm B: 27). The trial was stopped early due to insufficient accrual. The median overall survival (OS) rate was 11months [95% confidence interval: 9–20] and 8months [95%CI: 6–12], in the CG arm and in the C arm, respectively. The 1-year survival rate was 46% [95%CI: 28–64] in the combination arm and 35% [95%CI: 19–56] in the C arm (log rank test: p=0.73). The median progression-free survival (PFS) rate was 5 [95%CI: 3–11] and 3 [95%CI: 1–8] months in the CG and in the C arm, respectively. The 1-year PFS rate was 29% [95%CI: 15–48] in the combination arm and 15% [95%CI: 5–35] in the C arm (log rank test: p=0.27). No toxic deaths occurred. Grade 3–4 neutropenia (63% versus 12%) and grade 3–4 thrombocytopenia (37% versus 4%) were more frequent in the CG arm than in the C arm. ConclusionA non-significantly better outcome was observed with CG as compared to C in patients with CUP and a non-unfavourable prognosis. The toxicity profile of the combined arm was represented by haematologic toxicity with thrombocytopenia and leuconeutropenia. International collaboration is required to conduct phase III trials in patients with CUP.

Assessment of circulating tumor cells and serum markers for progression-free survival prediction in metastatic breast cancer: a prospective observational study.

IntroductionCirculating tumor cells (CTC) have been recently proposed as a new dynamic blood marker whose positivity at baseline is a prognostic factor and whose changes under treatment are correlated with progression-free survival (PFS) in metastatic breast cancer patients. However, serum marker levels are also used for the same purpose, and no clear comparison has been reported to date.MethodsThe IC 2006-04 enrolled prospectively 267 metastatic breast cancer patients treated by first line chemotherapy and confirmed that CTC levels are an independent prognostic factor for PFS and overall survival (OS). A secondary pre-planned endpoint was to compare prospectively the positivity rates and the value of CTC (CellSearch®), of serum tumor markers (carcinoembryonic antigen (CEA), cancer antigen 15.3 (CA 15-3), CYFRA 21-1), and of serum non-tumor markers (lactate deshydrogenase (LDH), alkaline phosphatase (ALP)) at baseline and under treatment for PFS prediction, independently from the other known prognostic factors, using univariate analyses and concordance indexes.ResultsA total of 90% of the patients had at least one elevated blood marker. Blood markers were correlated with poor performance status, high number of metastatic sites and with each other. In particular, CYFRA 21-1, a marker usually used in lung cancer, was elevated in 65% of patients. A total of 86% of patients had either CA 15-3 and/or CYFRA 21-1 elevated at baseline. Each serum marker was associated, when elevated at baseline, with a significantly shorter PFS. Serum marker changes during treatment, assessed either between baseline and week 3 or between baseline and weeks 6 to 9, were significantly associated with PFS, as reported for CTC. Concordance indexes comparison showed no clear superiority of any of the serum marker or CTC for PFS prediction.ConclusionsFor the purpose of PFS prediction by measuring blood marker changes during treatment, currently available blood-derived markers (CTC and serum markers) had globally similar performances. Besides CEA and CA 15-3, CYFRA 21-1 is commonly elevated in metastatic breast cancer and has a strong prognostic value.

Screening for Potential Covariates Influencing the Pharmacokinetics of Intravenous Busulfan: Results From a Large Pediatric Cohort Undergoing Hematopoietic Stem-Cell Transplantation

AbstractAbstract 1811 Background:Busulfan (Bu) is a standard component of HSCT regimens and shows a narrow therapeutic range. In very young children, Bu exhibits large pharmacokinetic (PK) variability. Intravenous (IV) Bu clearance was demonstrated to be non-linearly related to body weight (BW) and thus BW is used to optimally define the IV Bu dosage in children [Vassal G. et al., CCP 2006]. Search for additional relevant covariates is still a topical question to further explain the variability in young children (Pts) and infants. Objective:to screen for the effect of heterogeneous patient characteristics on IV Bu PK disposition, and to evaluate the consistency of weight-based dosing strategy (as recommended in the European SmPC) in a large cohort of children, especially in patients with rare diseases and a BW less than 9 kg. Patients and Method:The prospective observational study included 115 patients (pts), mostly infants (median age: 13 months [0.3-148], median BW: 9 kg [3.5-58]). Main diagnoses were immunodeficiency (n=71, mainly SCID and lymphohistiocytosis) and inherited metabolic disorders (n=15). Additional data (90 pts, mostly malignant diseases) from two prospective clinical trials previously described [Nguyen L. et al., BMT 2004] [Vassal G. et al., CCP 2006] were pooled with the observational dataset. Altogether, 205 children aged from 10 days to 15 years (median of 30 months) and weighed from 3.5 to 62.5 kg (median of 11.0 kg) were analysed. IV Bu (Busilvex®) was given over 2 h q6h × 16 doses. For most of the pts (88%), the starting doses were given according to the European SmPC: 1.0 mg/kg, 1.2 mg/kg, 1.1 mg/kg, 0.95 mg/kg and 0.8 mg/kg for pts <9 kg, 9 – <16 kg, 16–23 kg, >23–34 kg, and >34 kg BW, respectively.PK was assessed on plasma samples from the 1st, 9th and 13th dose (536 PK datasets). PK analysis was performed using Non Linear Mixed Effect modelling with the NONMEM program. Results:A one-compartment PK model suitably fitted the concentrations vs time data. BW remained the main determinant of IV Bu PK disposition. Increase of IV Bu clearance with child's growth was observed to be faster in younger than in older children. For children < 9 kg, a doubling in BW is associated with a 2.4-fold increase in Bu clearance while a 1.7-fold increase in Bu clearance is observed in pts ≥ 9 kg for the same BW growth. To take into account this difference, the relationships between Clearance (CL) and BW was modelled using a different allometric exponent for children with a BW of < 9 kg and ≥ 9 kg.The predictive equations were: CL (L/h) = 2.18 × (BW/9)1.25 and CL(L/h) = 2.18 × (BW/9)0.76 in pts < 9 kg and ≥ 9 kg, respectively. The inter-individual variability decreased from 64% (covariate-free model) to 23% for Bu CL. No further significant impact was identified even when considering large fluctuations of the following covariates: creatinine clearance (up to 11 × Upper Normal Limit), bilirubin (up to 4 × UNL), ferritine (up to 43 × UNL), hepatic transaminases (up to 39 × UNL), alkaline phosphatases (up to 4 × UNL), gGT (up to 86 × UNL), total protein (0.5 to 1.4 × UNL) and lactate deshydrogenase (up to 3 × UNL). No PK difference was observed between patients under Benzodiazepine and those under Phenytoin prophylaxis, neither when comparing different groups of diagnoses. Similar performances for targeting an AUC range of 900 – 1500 μM.min were achieved when comparing Bu dosing either adjusted according to the model predictive equation or with the use of the European SmPC recommendations. The success rates were about 60% and 80% in pts < 9 kg and ≥ 9 kg respectively. Conclusion:BW is confirmed to be the only relevant predictor to optimally define the IV Bu dosing in children. The population PK model has established a higher allometric exponent for younger children (< 9kg) in order to take into account the faster maturation of clearance in this weight group. This model is consistent with the dosing strategy recommended in the EMA registration. Disclosures:Daher Abdi:Pierre Fabre Médicament: Employment. Pétain:Pierre Fabre Médicament: Employment. Nguyen:Pierre Fabre Médicament: Employment.

Modulatory effect of fenugreek saponins on the activities of intestinal and hepatic disaccharidase and glycogen and liver function of diabetic rats

Diabetes mellitus is a serious health concern throughout the world and is often associated with a variety of bodily disorders such as liver toxicity and dysfunction. This study elucidates the effect of fenugreek saponin administration on disaccharidase and glycogen activities in the intestine and liver of surviving diabetic rats. It also evaluates the effect of saponin feeding using a number of liver toxicity indices, namely stress oxidant, liver dysfunction markers and metabolism. Our findings indicate that the fenugreek saponin fraction significantly modulated the disaccharidase and glycogen enzyme activities in the intestine and liver of rats, increased the hepatic glycogen content, suppressed the increase of blood glucose level and improved results in the oral glucose tolerance test (OGTT). The fenugreek saponin extract also efficiently protected the hepatic function, which was evidenced by the significant increases of superoxide dismutase (SOD), catalase (CAT), gluthation peroxidase (GPX), aspartate transaminase (AST), alanine transaminase (ALT) and lactate deshydrogenase (LDH) enzyme activities. Fenugreek saponin also induced a notable delay in the absorption of LDL-cholesterol and triglycerides and a remarkable increase in levels of HDL-cholesterol. A histological analysis of the hepatic tissues further established the positive effect of fenugreek saponin. Overall, the findings of the current study indicate that fenugreek saponins exhibit attractive properties and can be considered as promising candidates for future application as therapeutic agents in biotechnological and bioprocess-based technologies, particularly those related to the development of anti-diabetic, hepatoprotective and hypolipidemic drugs.

Field evaluation of a rapid diagnostic test (Parascreen™) for malaria diagnosis in the Peruvian Amazon

BackgroundThe rapid diagnostic tests for malaria (RDT) constitute a fast and opportune alternative for non-complicated malaria diagnosis in areas where microscopy is not available. The objective of this study was to validate a RDT named Parascreen™ under field conditions in Iquitos, department of Loreto, Peru. Parascreen™ is a RDT that detects the histidine-rich protein 2 (HRP2) antigen from Plasmodium falciparum and lactate deshydrogenase from all Plasmodium species.MethodsParascreen™ was compared with microscopy performed by experts (EM) and polymerase chain reaction (PCR) using the following indicators: sensitivity (Se), specificity (Sp), positive (PV+) and negative predictive values (PV-), positive (LR+) and negative likehood ratio (LR-).Results332 patients with suspected non-complicated malaria who attended to the MOH health centres were enrolled between October and December 2006. For P. falciparum malaria, Parascreen™ in comparison with EM, had Se: 53.5%, Sp: 98.7%, PV+: 66.7%, PV-: 97.8%, LR+: 42.27 and LR-: 0.47; and for non-P. falciparum malaria, Se: 77.1%, Sp: 97.6%, PV+: 91.4%, PV-: 92.7%, LR+: 32.0 and LR-: 0.22. The comparison of Parascreen™ with PCR showed, for P. falciparum malaria, Se: 81.8%, Sp: 99.1%, PV+: 75%, PV-: 99.4, LR+: 87.27 and LR-: 0.18; and for non-P. falciparum malaria Se: 76.1%, Sp: 99.2%, PV+: 97.1%, PV-: 92.0%, LR+: 92.51 and LR-: 0.24.ConclusionsThe study results indicate that Parascreen™ is not a valid and acceptable test for malaria diagnosis under the field conditions found in the Peruvian Amazon. The relative proportion of Plasmodium species, in addition to the genetic characteristics of the parasites in the area, must be considered before applying any RDT, especially after the finding of P. falciparum malaria parasites lacking pfhrp2 gene in this region.

Inhibitory effect of fenugreek galactomannan on digestive enzymes related to diabetes, hyperlipidemia, and liver-kidney dysfunctions

The present study was undertaken to investigate the effect of fenugreek galactomannan on intestinal glucose uptake in surviving diabetic rats. It explored their potential action with respect to lowering maltase, lactase, and sucrase activities in the small intestine of galactomannan-treated diabetic group compared to the diabetic control group. The findings indicate that the increase of blood glucose levels was significantly suppressed in the galactomannan-treated group than those in the diabetic rats. Moreover, the galactomannan isolated from fenugreek exhibited a prominent selective inhibitory effect against intestinal lipase activity. It was found to significantly delay the absorption of LDL-cholesterol and triglycerides and the increase in HDL-cholesterol. In addition, fenugreek galactomannan efficiently protect the hepatic function observed by the considerable decrease of aspartate and alanine transaminases (AST and ALT) and lactate deshydrogenase (LDH) contents in the serum of diabetic rats. The beneficial effects of fenugreek galactomannan were also evidenc-ed by their capacity to inhibit diabetes-induced kidney injury through lowering the urea and creatinine content in plasma. Overall, the conclusion of the present study indicate that fenugreek galactomannan displays a number of promising properties and attributes for future applications as therapeutic agents in biotechnological and bioprocess-based technologies, particularly those interested in the development of anti-diabetic and hypolipidemic drugs.

Iron nanoparticles increase 7-ketocholesterol-induced cell death, inflammation, and oxidation on murine cardiac HL1-NB cells

To evaluate the cytotoxicity of iron nanoparticles on cardiac cells and to determine whether they can modulate the biological activity of 7-ketocholesterol (7KC) involved in the development of cardiovascular diseases. Nanoparticles of iron labeled with Texas Red are introduced in cultures of nonbeating mouse cardiac cells (HL1-NB) with or without 7-ketocholesterol 7KC, and their ability to induce cell death, pro-inflammatory and oxidative effects are analyzed simultaneously. Flow cytometry (FCM), confocal laser scanning microscopy (CLSM), and subsequent factor analysis image processing (FAMIS) are used to characterize the action of iron nanoparticles and to define their cytotoxicity which is evaluated by enhanced permeability to SYTOX Green, and release of lactate deshydrogenase (LDH). Pro-inflammatory effects are estimated by ELISA in order to quantify IL-8 and MCP-1 secretions. Pro-oxidative effects are measured with hydroethydine (HE). Iron Texas Red nanoparticles accumulate at the cytoplasmic membrane level. They induce a slight LDH release, and have no inflammatory or oxidative effects. However, they enhance the cytotoxic, pro-inflammatory and oxidative effects of 7KC. The accumulation dynamics of SYTOX Green in cells is measured by CLSM to characterize the toxicity of nanoparticles. The emission spectra of SYTOX Green and nanoparticles are differentiated, and corresponding factor images specify the possible capture and cellular localization of nanoparticles in cells. The designed protocol makes it possible to show how Iron Texas Red nanoparticles are captured by cardiomyocytes. Interestingly, whereas these fluorescent iron nanoparticles have no cytotoxic, pro-inflammatory or oxidative activities, they enhance the side effects of 7KC.