Changes in aminoacidergic and monoaminergic neurotransmission in the hippocampus and amygdala of rats after ayahuasca ingestion

AIM: To evaluate changes in neurotransmission induced by a psychoactive beverage ayahuasca in the hippocampus and amygdala of naive rats. METHODS: The level of monoamines, their main metabolites and amino acid neurotransmitters concentrations were quantified using high performance liquid chromatography(HPLC). Four groups of rats were employed: saline-treated and rats receiving 250, 500 and 800 mg/kg of ayahuasca infusion(gavage). Animals were killed 40 min after drug ingestion and the structures stored at-80 ℃ until HPLC assay. The data from all groups were compared using Analysis of variance and Scheffé as post test and P 0.05 was accepted as significant. RESULTS: The results showed decreased concentrations of glycine(GLY)(0.13 ± 0.03 vs 0.29 ± 0.07, P 0.001) and γ-aminobutyric acid(GABA)(1.07 ± 0.14 vs 1.73 ± 0.25, P 0.001) in the amygdala of rats that received 500 of ayahuasca. Animals that ingested 800 mg/kg of ayahuasca also showed a reduction of GLY level(0.11 ± 0.01 vs 0.29 ± 0.07, P 0.001) and GABA(0.98 ± 0.06 vs 1.73 ± 0.25, P 0.001). In the hippocampus, increased GABA levels were found in rats that received all ayahuasca doses: 250 mg/kg(1.29 ± 0.19 vs 0.84 ± 0.21, P 0.05); 500 mg/kg(2.23 ± 038 vs 084 ± 0.21, P 0.05) and 800 mg/kg(1.98 ± 0.92 vs 0.84 ± 0.21, P 0.05). In addition, an increased utilization rate of all monoamines was found in the amygdala after ayahuasca administration in doses: 250 mg/kg(noradrenaline: 0.16 ± 0.02 vs 0.36 ± 0.06, P 0.01; dopamine: 0.39 ± 0.012 vs 2.39 ± 0.84, P 0.001; serotonin: 1.02 ± 0.22 vs 4.04 ± 0.91, P 0.001), 500 mg/kg(noradrenaline: 0.08 ± 0.02 vs 0.36 ± 0.06, P 0.001; dopamine: 0.33 ± 0.19 vs 2.39 ± 0.84, P 0.001; serotonin: 0.59 ± 0.08 vs 4.04 ± 0.91, P 0.001) and 800 mg/kg(noradrenaline: 0.16 ± 0.04 vs 0.36 ± 0.06, P 0.001; dopamine: 0.84 ± 0.65 vs2.39 ± 0.84, P 0.05; serotonin: 0.36 ± 0.02 vs 4.04 ± 0.91, P 0.001). CONCLUSION: Our data suggest increased release of inhibitory amino acids by the hippocampus and an increased utilization rate of monoamines by the amygdala after different doses of ayahuasca ingestion.     

“Immunonutrition” Has Failed to Improve Peritonitis-Induced Septic Shock in Rodents

Background

Immunonutrition in sepsis, including n-3 poly-unsaturated fatty acids (PUFAs) or L-arginine supplementation, is a controversial issue that has yielded a great number of studies for the last thirty-five years, and the conclusions regarding the quantity and quality of this support in patients are deceiving. The aim of the present experimental study is to investigate the effects of a pretreatment with enteral nutrition enriched with n-3 PUFAs or L-arginine on vascular dysfunctions, inflammation and oxidative stress during septic shock in rats.

Design

Rats were fed with enteral Peptamen^® HN (HN group), Peptamen^® AF containing n-3 PUFAs (AF group) or Peptamen^® AF enriched with L-arginine (AFA group). On day 4, peritonitis by cecal ligation and puncture (CLP) was performed. Rats were resuscitated (H18) once septic shock was established. After a 4-hour resuscitation, vessels and organs were harvested to assess inflammation, superoxide anion, nitric oxide and prostacyclin levels. Ex-vivo vascular reactivity was also performed.

Results

Compared to CLP-AF or CLP-HN groups, 47.6% of CLP-AFA rats died before the beginning of hemodynamic measurements (vs. 8.0% and 20.0% respectively, p<0.05). AF and AFA rats required significantly increased norepinephrine infusion rates to reach the mean arterial pressure objective, compared to CLP-HN rats. Both CLP-AF and CLP-AFA reduced mesenteric resistance arterial contractility, decreased vascular oxidative stress, but increased NF-κB (0.40±0.15 in CLP-AF and 0.69±0.06 in CLP-AFA vs. 0.09±0.03 in SHAM rats and 0.30±0.06 in CLP-HN, ß-actin ratio, p<0.05) and pIκB expression (0.60±0.03 in CLP-AF and 0.94±0.15 in CLP-AFA vs. 0.04±0.01 in SHAM rats and 0.56±0.07 in CLP-HN, ß-actin ratio, p<0.05), nitric oxide and prostacyclin production in septic rats.

Conclusions

Although n-3 PUFAs or L-arginine supplementation exhibited an antioxidant effect, it worsened the septic shock-induced vascular dysfunction. Furthermore, mortality was higher after L-arginine supplementation.     

Updates in the pathophysiological mechanisms of Parkinson's disease: Emerging role of bone marrow mesenchymal stem cells

AIM: To explore the approaches exerted by mesenchymal stem cells(MSCs) to improve Parkinson's disease(PD) pathophysiology.METHODS: MSCs were harvested from bone marrowof femoral bones of male rats, grown and propagated in culture. Twenty four ovariectomized animals were classified into 3 groups: Group(1) was control, Groups(2) and(3) were subcutaneously administered with rotenone for 14 d after one month of ovariectomy for induction of PD. Then, Group(2) was left untreated, while Group(3) was treated with single intravenous dose of bone marrow derived MSCs(BM-MSCs). SRY gene was assessed by PCR in brain tissue of the female rats. Serum transforming growth factor beta-1(TGF-β1), monocyte chemoattractant protein-1(MCP-1) and brain derived neurotrophic factor(BDNF) levels were assayed by ELISA. Brain dopamine DA level was assayed fluorometrically, while brain tyrosine hydroxylase(TH) and nestin gene expression were detected by semi-quantitative real time PCR. Brain survivin expression was determined by immunohistochemical procedure. Histopathological investigation of brain tissues was also done.RESULTS: BM-MSCs were able to home at the injured brains and elicited significant decrease in serum TGF-β1(489.7 ± 13.0 vs 691.2 ± 8.0, P 0.05) and MCP-1(89.6 ± 2.0 vs 112.1 ± 1.9, P 0.05) levels associated with significant increase in serum BDNF(3663 ± 17.8 vs 2905 ± 72.9, P 0.05) and brain DA(874 ± 15.0 vs 599 ± 9.8, P 0.05) levels as well as brain TH(1.18 ± 0.004 vs 0.54 ± 0.009, P 0.05) and nestin(1.29 ± 0.005 vs 0.67 ± 0.006, P 0.05) genes expression levels. In addition to, producing insignificant increase in the number of positive cells for survivin(293.2 ± 15.9 vs 271.5 ± 15.9, P 0.05) expression. Finally, the brain sections showed intact histological structure of the striatum as a result of treatment with BM-MSCs. CONCLUSION: The current study sheds light on the therapeutic potential of BM-MSCs against PD pathophysiology via multi-mechanistic actions.     

Antenatal Dexamethasone after Asphyxia Increases Neural Injury in Preterm Fetal Sheep

Background and Purpose

Maternal glucocorticoid treatment for threatened premature delivery dramatically improves neonatal survival and short-term morbidity; however, its effects on neurodevelopmental outcome are variable. We investigated the effect of maternal glucocorticoid exposure after acute asphyxia on injury in the preterm brain.

Methods

Chronically instrumented singleton fetal sheep at 0.7 of gestation received asphyxia induced by complete umbilical cord occlusion for 25 minutes. 15 minutes after release of occlusion, ewes received a 3 ml i.m. injection of either dexamethasone (12 mg, n = 10) or saline (n = 10). Sheep were killed after 7 days recovery; survival of neurons in the hippocampus and basal ganglia, and oligodendrocytes in periventricular white matter were assessed using an unbiased stereological approach.

Results

Maternal dexamethasone after asphyxia was associated with more severe loss of neurons in the hippocampus (CA3 regions, 290±76 vs 484±98 neurons/mm², mean±SEM, P<0.05) and basal ganglia (putamen, 538±112 vs 814±34 neurons/mm², P<0.05) compared to asphyxia-saline, and with greater loss of both total (913±77 vs 1201±75/mm², P<0.05) and immature/mature myelinating oligodendrocytes in periventricular white matter (66±8 vs 114±12/mm², P<0.05, vs sham controls 165±10/mm², P<0.001). This was associated with transient hyperglycemia (peak 3.5±0.2 vs. 1.4±0.2 mmol/L at 6 h, P<0.05) and reduced suppression of EEG power in the first 24 h after occlusion (maximum −1.5±1.2 dB vs. −5.0±1.4 dB in saline controls, P<0.01), but later onset and fewer overt seizures.

Conclusions

In preterm fetal sheep, exposure to maternal dexamethasone during recovery from asphyxia exacerbated brain damage.     

Increased Short-Term Beat-To-Beat Variability of QT Interval in Patients with Acromegaly

Cardiovascular diseases, including ventricular arrhythmias are responsible for increased mortality in patients with acromegaly. Acromegaly may cause repolarization abnormalities such as QT prolongation and impairment of repolarization reserve enhancing liability to arrhythmia. The aim of this study was to determine the short-term beat-to-beat QT variability in patients with acromegaly. Thirty acromegalic patients (23 women and 7 men, mean age±SD: 55.7±10.4 years) were compared with age- and sex-matched volunteers (mean age 51.3±7.6 years). Cardiac repolarization parameters including frequency corrected QT interval, PQ and QRS intervals, duration of terminal part of T waves (T_peak-T_end) and short-term variability of QT interval were evaluated. All acromegalic patients and controls underwent transthoracic echocardiographic examination. Autonomic function was assessed by means of five standard cardiovascular reflex tests. Comparison of the two groups revealed no significant differences in the conventional ECG parameters of repolarization (QT: 401.1±30.6 ms vs 389.3±16.5 ms, corrected QT interval: 430.1±18.6 ms vs 425.6±17.3 ms, QT dispersion: 38.2±13.2 ms vs 36.6±10.2 ms; acromegaly vs control, respectively). However, short-term beat-to-beat QT variability was significantly increased in acromegalic patients (4.23±1.03 ms vs 3.02±0.80, P<0.0001). There were significant differences between the two groups in the echocardiographic dimensions (left ventricular end diastolic diameter: 52.6±5.4 mm vs 48.0±3.9 mm, left ventricular end systolic diameter: 32.3±5.2 mm vs 29.1±4.4 mm, interventricular septum: 11.1±2.2 mm vs 8.8±0.7 mm, posterior wall of left ventricle: 10.8±1.4 mm vs 8.9±0.7 mm, P<0.05, respectively). Short-term beat-to-beat QT variability was elevated in patients with acromegaly in spite of unchanged conventional parameters of ventricular repolarization. This enhanced temporal QT variability may be an early indicator of increased liability to arrhythmia.     

Long-Term Treatment of Clonidine,Atenolol, Amlodipine and Dihydrochlorothiazide,but Not Enalapril,Impairs the Sexual Function in Male Spontaneously Hypertensive Rats

This study was designed to investigate the impact of representative antihypertensive drugs of 5 classes on the sexual function in male spontaneously hypertensive rats (SHR) at doses that achieved similar blood pressure (BP) reduction. The experiment was performed in 6 groups of male SHR. The dose are 20 μg/kg/day for clonidine, 3 mg/kg/day for enalapril, 20 mg/kg/day for atenolol, 2 mg/kg/day for amlodipine, and 10 mg/kg/day for dihydrochlorothiazide. SHR were treated for 3 months, and then the penile erection and sexual behavior were detected. After BP recording, SHR were killed to evaluate the organ-damage, weight of accessory sex organs and levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and testosterone in serum. Five drugs had the similar efficacy on BP reduction. All drugs except of enalapril, significantly prolonged the mount latency, and decreased the mount frequency (P<0.05). Clonidine also reduced the conception rate (45% vs. 80% in control group, P<0.05). Amlodipine and dihydrochlorothiazide significantly increased the testosterone level (0.79±0.30, 0.80±0.34 vs. 0.49±0.20 in control group, unit: ng/dl, P<0.05). Enalapril, atenolol and amlodipine also significantly decreased the BP variability (systolic, 8.2±2.5, 7.6±1.8, 8.9±2.0 vs. 12.2±3.8 in control group, unit: mm Hg). All these drugs significantly decreased the organ-damage (P<0.05). In conclusion, long-term treatment with 5 common antihypertensive drugs possessed obvious organ protection in SHR. Clonidine, atenolol, amlodipine and dihydrochlorothiazide, but not enalapril, impair sexual function.     

Kinetics of Label Retaining Cells in the Developing Rat Kidneys

Background

The kidney is a specialized low-regenerative organ with several different types of cellular lineages. The BrdU label-retaining cell (LRCs) approach has been used as part of a strategy to identify tissue-specific stem cells in the kidney; however, because the complementary base pairing in double-stranded DNA blocks the access of the anti-BrdU antibody to BrdU subunits, the stem cell marker expression in BrdU-labeled cells are often difficult to detect. In this study, we introduced a new cell labeling and detection method in which BrdU was replaced with 5-ethynyl-2-deoxyuridine (EdU) and examined the time-dependent dynamic changes of EdU-labeled cells and potential stem/progenitor markers in the development of kidney.

Methods

Newborn rats were intraperitoneally injected with EdU, and their kidneys were harvested respectively at different time points at 1 day, 3 days, 1 week, 2 weeks, and 6 weeks post-injection. The kidney tissues were processed for EdU and cellular markers by immunofluorescence staining.

Results

At the early stage, LRCs labeled by EdU were 2176.0 ± 355.6 cells at day one in each renal tissue section, but dropped to 168 ± 48.4 cells by week 6. As time increased, the numbers of LRCs were differentially expressed in the renal cortex and papilla. At the postnatal day one, nearly twice as many cells in the cortex were EdU-labeled as compared to the papilla (28.6 ± 3.6% vs. 15.6 ± 3.4%, P<0.05), while there were more LRCs within the renal papilla since the postnatal week one, and at the postnatal week 6, one third as many cells in the cortex were EdU-labeled as compared to the papilla (2.5 ± 0.1% vs. 7.7 ± 2.7%, P<0.05). The long-term LRCs at 6-week time point were associated exclusively with the glomeruli in the cortex and the renal tubules in the papilla. At 6 weeks, the EdU-labeled LRCs combined with expression of CD34, RECA-1, Nestin, and Synaptopodin were discretely but widely distributed within the glomeruli; Stro-1 around the glomeruli; and α-smooth muscle actin (SMA) in arteries. Conversely, co-expression of CD34, RECA-1, and Nestin with the long term EdU-labeled LRCs was significantly lower in renal tubules (P<0.01), while Stro-1 and Synaptopodin were not detected.

Conclusion

Our data found that at 6-week time point, EdU-labeled LRCs existing in the glomeruli expressed undifferentiated podocyte and endothelial markers at high rates, while those in the renal tubules expressed Nestin and vascular markers at low rates. To understand the characterization and localization of these EdU-LRCs, further studies will be needed to test cell lineage tracing, clonogenicity and differentiation potency, and the contributions to the regeneration of the kidney in response to renal injury/repair.     

Experimental Type 2 Diabetes Induces Enzymatic Changes in Isolated Rat Enterocytes

Diabetes in humans and in experimental animals produces changes in the function and structure of the small intestine. The authors determined the activity of intestinal disaccharidases (maltase and sucrase) and of 6-phosphofructo-1-kinase (PFK-1) in enterocytes isolated from the small intestine of male Wistar rats (2.5 to 3 months old) with experimental nonobese type 2 diabetes, induced by streptozotocin (STZ) injection on the day of birth (n0-STZ) or on the 5th day of life (n5-STZ), with different degrees of hyperglycemia and insulinemia (n0-STZ and n5-STZ models). The glycemia (mmol/L) of the diabetic rats (n0-STZ: 8.77 ± 0.47; n5-STZ: 20.83 ± 0.63) was higher (P < .01) than that of the nondiabetic (ND) rats (5.99 ± 0.63); on the contrary, the insulinemia (ng/mL) was significantly lower in both n0-STZ (1.74 ± 0.53; P < .05) and n5-STZ (1.12 ± 0.44; P < .01) diabetic rats than in normal rats (3.77 ± 0.22). The sucrase and maltase activities (U/g protein) in diabetic rats (n0-STZ: 89 ± 9 and 266 ± 12; n5-STZ: 142 ± 23 and 451 ± 57) were significantly higher than those in the ND group (66 ± 5 and 228 ± 22). The PFK-1 activities (mU/mg protein) in the diabetic models (n0-STZ: 14.89 ± 1.51; n5-STZ: 13.35 ± 3.12) were significantly lower (P < .05) than in ND rats (20.54 ± 2.83). The data demonstrated enzymatic alterations in enterocytes isolated fromthe small intestine of n0-STZ rats that are greater (P < .05) than in the more hyperglycemic and hypoinsulinemic n5-STZ animals. The results also show that nonobese type 2–like diabetes in the rat produces modifications that favor an increase in glucose absorption rates.     

Suppression of Experimental Choroidal Neovascularization by Curcumin in Mice

Purpose

To investigate the effects of curcumin on the development of experimental choroidal neovascularization (CNV) with underlying cellular and molecular mechanisms.

Methods

C57BL/6N mice were pretreated with intraperitoneal injections of curcumin daily for 3 days prior to laser-induced CNV, and the drug treatments were continued until the end of the study. The CNV area was analyzed by fluorescein-labeled dextran angiography of retinal pigment epithelium (RPE)-choroid flat mounts on day 7 and 14, and CNV leakage was evaluated by fluorescein angiography (FA) on day 14 after laser photocoagulation. The infiltration of F4/80 positive macrophages and GR-1 positive granulocytes were evaluated by immunohistochemistry on RPE-choroid flat mounts on day 3. Their expression in RPE-choroid complex was quantified by real-time PCR (F4/80) and Western blotting (GR-1) on day 3. RPE-choroid levels of vascular endothelial growth factor (VEGF), tumor necrosis factor (TNF)-α, monocyte chemotactic protein (MCP)-1, and intercellular adhesion molecule (ICAM)-1 were examined by ELISA on day 3. Double immunostaining of F4/80 and VEGF was performed on cryo-sections of CNV lesions on day 3. The expression of nuclear factor (NF)-κB and hypoxia-inducible factor (HIF)−1α in the RPE-choroid was determined by Western blotting.

Results

Curcumin-treated mice had significantly less CNV area (P<0.05) and CNV leakage (P<0.001) than vehicle-treated mice. Curcumin treatment led to significant inhibition of F4/80 positive macrophages (P<0.05) and GR-1 positive granulocytes infiltration (P<0.05). VEGF mainly expressed in F4/80 positive macrophages in laser injury sites, which was suppressed by curcumin treatment (P<0.01). Curcumin inhibited the RPE-choroid levels of TNF-α (P<0.05), MCP-1 (P<0.05) and ICAM-1 (P<0.05), and suppressed the activation of NF-κB in nuclear extracts (P<0.05) and the activation of HIF−1α (P<0.05).

Conclusion

Curcumin treatment led to the suppression of CNV development together with inflammatory and angiogenic processes including NF-κB and HIF−1α activation, the up-regulation of inflammatory and angiogenic cytokines, and infiltrating macrophages and granulocytes. This provides molecular and cellular evidence of the validity of curcumin supplementation as a therapeutic strategy for the suppression of age-related macular degeneration (AMD)-associated CNV.     

Infusion of Trx-1-Overexpressing hucMSC Prolongs the Survival of Acutely Irradiated NOD/SCID Mice by Decreasing Excessive Inflammatory Injury

A protective reagent for ARI should have the ability to repair injured tissue caused by radiation and prevent continuous damage from secondary risk factors. Trx-1 was explored as a candidate therapy for ARI, as it scavenges reactive oxygen species, regulates cell growth and differentiation, participates in immune reactions, and inhibits apoptosis by acting inside and/or outside cells. Trx-1 can also decrease excessive inflammation in ARI by regulating the creation of inflamed media, by inhibiting the activation of complement, and by reducing the chemotaxis, adhesion, and migration of inflammatory cells. As effectively and stably expressing exogenous genes in the long term and regulating immune inflammation and tissue repair, MSC are a good choice for Trx-1 gene therapy. In this study, Trx-1-overexpressing hucMSC-Trx-1 were obtained by adenoviral vector-mediated infection. We first measured the redox capacity of hucMSC-Trx-1 with an antioxidant capacity (T-AOC) assay, a hydrogen peroxide (H₂O₂) content determination assay in vivo, a H₂O₂-induced oxidation hemolysis assay, and a lipid peroxidation assay in vitro. Then, we measured survival time, the protection of the hematopoietic system, and the regulation of inflammation in important organs in three treatment groups of NOD/SCID mice (treated with hucMSC-Trx-1, with hucMSC, and with saline) that were exposed to 4.5 Gy ⁶⁰Co-γ-ray radiation. The hucMSC-Trx-1 group achieved superior antioxidation results, protecting bone marrow hematopoietic stem cells (Lin⁻CD117⁺: hucMSC-Trx-1 vs. hucMSC, P<0.05; hucMSC-Trx-1 vs. NS, P<0.01), promoting the formation of red blood cells and hemoglobin (hucMSC-Trx-1 vs. hucMSC or NS, P<0.05), reducing inflammation and damage in important organs (Bone marrow and lung: hucMSC-Trx-1 vs. NS, P<0.01; hucMSC-Trx-1 vs. hucMSC, P<0.05. Liver and intestine: hucMSC-Trx-1 vs. NS, P<0.05; hucMSC-Trx-1 vs. hucMSC, P<0.05), and prolonging survival (hucMSC-Trx-1 vs. hucMSC or NS, P<0.01). Therefore, hucMSC-Trx-1 combines the merits of gene and cell therapy as a multifunctional radioprotector for ARI.     

In Vitro and In Vivo Investigation of the Angiogenic Effects of Liraglutide during Islet Transplantation

Introduction

This study investigated the angiogenic properties of liraglutide in vitro and in vivo and the mechanisms involved, with a focus on Hypoxia Inducible Factor-1α (HIF-1α) and mammalian target of rapamycin (mTOR).

Materials and Methods

Rat pancreatic islets were incubated in vitro with 10 μmol/L of liraglutide (Lira) for 12, 24 and 48 h. Islet viability was studied by fluorescein diacetate/propidium iodide staining and their function was assessed by glucose stimulation. The angiogenic effect of liraglutide was determined in vitro by the measure of vascular endothelial growth factor (VEGF) secretion using enzyme-linked immunosorbent assay and by the evaluation of VEGF and platelet-derived growth factor-α (PDGFα) expression with quantitative polymerase chain reaction technic. Then, in vitro and in vivo, angiogenic property of Lira was evaluated using immunofluorescence staining targeting the cluster of differentiation 31 (CD31). To understand angiogenic mechanisms involved by Lira, HIF-1α and mTOR activation were studied using western blotting. In vivo, islets (1000/kg body-weight) were transplanted into diabetic (streptozotocin) Lewis rats. Metabolic control was assessed for 1 month by measuring body-weight gain and fasting blood glucose.

Results

Islet viability and function were respectively preserved and enhanced (p<0.05) with Lira, versus control. Lira increased CD31-positive cells, expression of VEGF and PDGFα (p<0.05) after 24 h in culture. Increased VEGF secretion versus control was also observed at 48 h (p<0.05). Moreover, Lira activated mTOR (p<0.05) signalling pathway. In vivo, Lira improved vascular density (p<0.01), body-weight gain (p<0.01) and reduced fasting blood glucose in transplanted rats (p<0.001).

Conclusion

The beneficial effects of liraglutide on islets appeared to be linked to its angiogenic properties. These findings indicated that glucagon-like peptide-1 analogues could be used to improve transplanted islet revascularisation.     

Contribution of Baicalin on the Plasma Protein Binding Displacement and CYP3A Activity Inhibition to the Pharmacokinetic Changes of Nifedipine in Rats In Vivo and In Vitro

Baicalin purified from the root of Radix scutellariae is widely used in clinical practices. This study aimed to evaluate the effect of baicalin on the pharmacokinetics of nifedipine, a CYP3A probe substrate, in rats in vivo and in vitro. In a randomised, three-period crossover study, significant changes in the pharmacokinetics of nifedipine (2 mg/kg) were observed after treatment with a low (0.225 g/kg) or high (0.45 g/kg) dose of baicalin in rats. In the low- and high-dose groups of baicalin-treated rats, C _max of total nifedipine decreased by 40%±14% (P<0.01) and 65%±14% (P<0.01), AUC_0–∞ decreased by 41%±8% (P<0.01) and 63%±7% (P<0.01), V_d increased by 85%±43% (P<0.01) and 224%±231% (P<0.01), and CL increased by 97%±78% (P<0.01) and 242%±135% (P<0.01), respectively. Plasma protein binding experiments in vivo showed that C _max of unbound nifedipine significantly increased by 25%±19% (P<0.01) and 44%±29% (P<0.01), respectively, and there was a good correlation between the unbound nifedipine (%) and baicalin concentrations (P<0.01). Furthermore, in vitro results revealed that baicalin was a competitive displacer of nifedipine from plasma proteins. In vitro incubation experiments demonstrated that baicalin could also competitively inhibit CYP3A activity in rat liver microsomes in a concentration-dependent manner. In conclusion, the pharmacokinetic changes of nifedipine may be modulated by the inhibitory effects of baicalin on plasma protein binding and CYP3A–mediated metabolism.     

Microsporidia and Its Relation to Crohn's Disease. A Retrospective Study

Background

The cause of Crohn''s Disease (CD) remains unknown. Recently a decrease in the global lymphocyte population in the peripheral blood of CD patients has been reported. This decrease was more evident in γδ T lymphocytes, especially γδ CD8+T subsets. Furthermore, a decrease of IL-7 was also observed in these patients. We propose the hypothesis that microsporidia, an obligate intracellular opportunistic parasite recently related to fungi, in CD patients can take advantage of the lymphocytes and IL-7 deficits to proliferate and to contribute to the pathophysiology of this disease.

Methods and Findings

In this case-control study, serum samples were collected from 36 CD patients and from 36 healthy individuals (controls), IgE and IgG anti-Encephalitozoon antibodies were determined by ELISA; and forty-four intestinal tissue samples were analyzed through real time Polymerase Chain Reaction (PCR), twenty CD patients, nine with others diseases and 15 healthy subjects.We observed that IgE anti-Encephalitozoon levels were significantly higher in patients with CD: 0.386(±0.256) vs control group, 0.201(±0.147), P<0.001. However, IgG anti-Encephalitozoon values were significantly lower in CD patients: 0.361(±0.256) vs control group, 0.876(±0.380), P<0.001. In the group of CD patients, 6/20 (30%) were positive by real time PCR for microsporidia and, all the patients of the control group were negative by real time PCR.

Conclusions

These results suggest that CD patients are a group at risk for microsporidiasis and, moreover that microsporidia may be involved as a possible etiologic factor of CD.     

Ocular Dominance Is Associated with the Ganglion Cell-Inner Plexiform Layer Thickness Profile in the Macula

Purpose

To investigate the characteristics of macular ganglion cell-inner plexiform layer (GCIPL) thickness profiles associated with ocular dominance.

Setting

Private practice, Seoul, Republic of Korea.

Design

Comparative case-control study.

Methods

Both eyes of 199 participants with no ophthalmic abnormalities were included. Participants were imaged by spectral-domain optical coherence tomography, and underwent dominant eye testing using a hole-in-a-card test (sighting dominance) at the same visit. Macular GCIPL, as well as circumpapillary retinal nerve fiber layer (RNFL) thickness were compared for individual patients, according to ocular dominance.

Results

Ocular dominance occurred predominantly in the right eye (right vs. left: 72.36 vs. 27.60%; P < 0.001). In the comparison of macular GCIPL thickness, the average (81.27±5.01 μm vs. 80.66±6.31 μm in dominant vs. non-dominant eyes), inferonasal (81.39±5.47μm vs. 80.33±6.82μm, and inferior sectors (77.95±6.05μm vs. 76.97±8.15μm) were significantly different between dominant and non-dominant eyes (P = 0.040, 0.005, and 0.032, respectively). Significant predictors of average GCIPL thickness were spherical equivalent (β = 1.37, P<0.001), astigmatic power (β = 1.44, P = 0.009), disc area (β = 3.90, P < 0.001), average RNFL thickness (β = 0.22, P<0.001), average cup-to-disc ratio (β = 5.74, P = 0.002), difference between the inferior and superior quadrant RNFL thicknesses (β = 0.08, P = 0.024), and ocular dominance (β = 2.10, P = 0.020). On multivariate regression analysis, ocular dominance was correlated with average GCIPL thickness after adjusting for potential confounders (β = 1.63, P = 0.048).

Conclusions

Dominant eyes accompanied significantly thicker average macular GCIPL. This information suggests that macular GCIPL thickness may provide an indicator of the relative dominance of an eye.     

Ovariectomy differential influence on some hemostatic markers of mice and rats

Rodent ovariectomy is an experimental method to eliminate the main source of sexual steroids. This work explored for the first time the ovariectomy temporal changes induced in the hemostatic coagulation markers: prothrombin time (PT), activated partial thromboplastin time (aPTT), thrombin time (TT), and fibrinogen concentration (FIB) along with uterine weight on adult female CD1 mice and Wistar rats. Uterine weight (Uw) was assessed before ovariectomy (control), and 1, 3, 5, 7, 9, 16, and 21 days after surgery. PT, aPTT, TT and FIB were estimated the same days, using reported standard techniques. Ovariectomy decreased Uw, since day 1; and from day 10 to 21 reached the lowest values for both species. After day 1, mice hemostatic parameters changed (PT +10%, P<0.05; aPTT +53%, P<0.05; TT −24%, P<0.05; FIB +67%, P<0.05). Rats showed significant changes only in TT and FIB (TT −13%, P<0.001; FIB +65%, P<0.001). Neither mice PT, aPTT and TT, recovered control values after 21 days. In the rats from day 5 to 16 aPTT diminished (18–23%, P<0.05) recovering to control values on day 21, TT after 9 days and PT on day 16. In both species, FIB returned to its control values after 9 days. Ovariectomy differentially altered the PT hemostatic parameter of mice and rats indicating a non-equivalence among both species behaviour for experimental studies of blood coagulation.     

A Pro-Inflammatory Role of C5L2 in C5a-Primed Neutrophils for ANCA-Induced Activation

Background

The complement system is crucial for the development of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). In particular, C5a and its receptor on neutrophils, CD88, play a central role. The functional role of the second receptor of C5a, C5L2, remains unclear. In the current study, we investigated the role of C5L2 in C5a-primed neutrophils for ANCA-induced activation.

Methods

The effect of blocking C5L2 by anti-human C5L2 blocking antibody were tested on respiratory burst and degranulation of C5a-primed neutrophils activated with ANCA, as well as on membrane-bound proteinase 3 (mPR3) and concentration of myeloperoxidase (MPO) in supernatant of C5a-primed neutrophils. An antagonist for CD88 was also employed.

Results

Blocking C5L2 resulted in a significantly decreased MPO concentration in the supernatant of C5a-primed neutrophils. mPR3 expression increased from 209.0±43.0 in untreated cells to 444.3±60.8 after C5a treatment (P<0.001), and decreased to 375.8±65.44, 342.2±54.3 and 313.7±43.6 by pre-incubating blocking C5L2 antibody at 2.5 µg/ml, 5 µg/ml or 10 µg/ml (compared with C5a-priming group, P<0.001, P<0.001, and P<0.001), respectively. In C5a-primed neutrophils, subsequently activating with MPO-ANCA-positive IgG, the MFI value was 425.8±160.6, which decreased to 292.8±141.2, 289.7±130.0 and 280.3±136.4 upon pre-incubation with mouse anti-human C5L2 blocking antibody at 2.5 µg/ml, 5 µg/ml or 10 µg/ml (compared with C5a-primed neutrophils, for MPO-ANCA-positive IgG-induced activation, P<0.05, P<0.05, and P<0.05), respectively. Blocking C5L2 also resulted in significantly decreased C5a-primed neutrophils for PR3-ANCA-positive IgG-induced activation. Moreover, the lactoferrin concentration in the supernant significantly decreased in pre-incubation with anti-human C5L2 blocking antibody, compared with C5a-primed neutrophils induced by PR3- or MPO-ANCA-positive IgG.

Conclusions

C5L2 may be implicated in the pro-inflammatory role in C5a-primed neutrophils for ANCA-induced activation.     

Mesangial Cell-Binding Activity of Serum Immunoglobulin G in Patients with Lupus Nephritis

In vitro data showed that immunoglobulin G (IgG) from patients with lupus nephritis (LN) could bind to cultured human mesangial cells (HMC). The clinical relevance of such binding was unknown. Binding of IgG and subclasses was measured in 189 serial serum samples from 23 patients with Class III/IV±V LN (48 during renal flares, 141 during low level disease activity (LLDA)). 64 patients with non-lupus glomerular diseases (NLGD) and 23 healthy individuals were used as controls. HMC-binding was measured with cellular ELISA and expressed as OD index. HMC-binding index of total IgG was 0.12±0.09, 0.36±0.25, 0.59±0.37 and 0.74±0.42 in healthy subjects, NLGD, LN patients during LLDA, and LN flares respectively (P = 0.046, LN flare vs. LLDA; P<0.001, for healthy controls or NLGD vs. LN during flare or LLDA). Binding of serum IgG₁ to HMC was 0.05±0.05, 0.15±0.11, 0.41±0.38 and 0.55±0.40 for the corresponding groups respectively (P = 0.007, LN flare vs. remission; P<0.001, for healthy controls or NLGD vs. LN during flare or remission). IgG₂, IgG₃ and IgG₄ from patients and controls did not show significant binding to HMC. Total IgG and IgG₁ HMC-binding index correlated with anti-dsDNA level (r = 0.26 and 0.39 respectively, P<0.001 for both), and inversely with C3 (r = −0.17 and −0.45, P<0.05 for both). Sensitivity/specificity of total IgG or IgG₁ binding to HMC in predicting renal flares were 81.3%/39.7% (ROC AUC 0.61, P = 0.03) and 83.8%/41.8% (AUC 0.63, P = 0.009) respectively. HMC-binding by IgG_1, but not total IgG, correlated with mesangial immune deposition in LN renal biopsies under electron microscopy. Our results showed that binding of serum total IgG and IgG₁ in LN patients correlates with disease activity. The correlation between IgG₁ HMC-binding and mesangial immune deposition suggests a potential pathogenic significance.