Effect of Various Pathological Conditions on Nitric Oxide Level and L-Citrulline Uptake in Motor Neuron-Like (NSC-34) Cell Lines.

Amyotrophic lateral sclerosis (ALS) is a fatal motor neuron disorder that causes progressive paralysis. L-Citrulline is a non-essential neutral amino acid produced by L-arginine via nitric oxide synthase (NOS). According to previous studies, the pathogenesis of ALS entails glutamate toxicity, oxidative stress, protein misfolding, and neurofilament disruption. In addition, L-citrulline prevents neuronal cell death in brain ischemia; therefore, we investigated the change in the transport of L-citrulline under various pathological conditions in a cell line model of ALS. We examined the uptake of [14C]L-citrulline in wild-type (hSOD1wt/WT) and mutant NSC-34/ SOD1G93A (MT) cell lines. The cell viability was determined via MTT assay. A transport study was performed to determine the uptake of [14C]L-citrulline. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis was performed to determine the expression levels of rat large neutral amino acid transported 1 (rLAT1) in ALS cell lines. Nitric oxide (NO) assay was performed using Griess reagent. L-Citrulline had a restorative effect on glutamate induced cell death, and increased [14C]L-citrulline uptake and mRNA levels of the large neutral amino acid transporter (LAT1) in the glutamate-treated ALS disease model (MT). NO levels increased significantly when MT cells were pretreated with glutamate for 24 h and restored by co-treatment with L-citrulline. Co-treatment of MT cells with L-arginine, an NO donor, increased NO levels. NSC-34 cells exposed to high glucose conditions showed a significant increase in [14C]L-citrulline uptake and LAT1 mRNA expression levels, which were restored to normal levels upon co-treatment with unlabeled L-citrulline. In contrast, exposure of the MT cell line to tumor necrosis factor alpha, lipopolysaccharides, and hypertonic condition decreased the uptake significantly which was restored to the normal level by co-treating with unlabeled L-citrulline. L-Citrulline can restore NO levels and cellular uptake in ALS-affected cells with glutamate cytotoxicity, pro-inflammatory cytokines, or other pathological states, suggesting that L-citrulline supplementation in ALS may play a key role in providing neuroprotection.

Differences in Neurotransmitters Level as Biomarker on Sleep Effects in Dementia Patients with Insomnia after Essential Oils Treatment.

This study aimed to examine the effects of several essential oils on insomnia in dementia patients following transdermal treatment (aromatherapy). The mean change rates (%) of sleep biomarkers were compared between the single essential and jojoba (vehicle) oil massage groups in this study. The lavender (L) essential oil massage group demonstrated a significant decrease in the mean change rate (%) of 24-h urinary free cortisol, whereas the valerian (V) essential oil massage group demonstrated a significant increase in the mean change rate (%) of serum 5-hydroxytryptamine. In addition, a significant increase in the mean change rate (%) of 24-h urinary norepinephrine was observed in the chamomile (C) essential oil massage group only. Based on these results, valerian, lavender, and chamomile oils were mixed in different ratios to produce blending oils A (L:C:V=2:2:1), B (L:C:V=3:1:1) and C (L:C:V=1:3:1). The highest level of serum 5-hydroxytryptamine was observed after administering blending oil A. These results suggest that blending oil A might possess therapeutic effects against insomnia. Overall, it is hypothesized that the optimally blended essential oil will produce synergic effects when combined with hypnotic drugs.

Cognitive behavior therapy for diabetes distress, depression, health anxiety, quality of life and treatment adherence among patients with type-II diabetes mellitus: a randomized control trial.

OBJECTIVE: Diabetes distress typically causes depressive symptoms; common comorbidity of diabetes unpleasantly affects patients' medical and psychological functions. Psychotherapeutic interventions are effective treatments to treat depressive symptoms and to improve the quality of life in many chronic diseases including diabetes. The present study investigated the efficacy of cognitive behavior therapy (CBT) to treat depressive symptoms in patients with type 2 diabetes mellitus (T2DM) using experimental and waitlist control conditions. MATERIALS AND METHODS: A total of 130 diagnosed patients with T2DM were taken from outdoor patients services of different hospitals in Faisalabad. Ninety patients met the eligibility criteria and were randomly assigned to experimental (n = 45) and waitlist control (n = 45) conditions. All the patients completed clinical interviews and assessment measures at pre-and post-assessment stages (16 weeks intervals). Medical consultants at the respective hospitals diagnosed the patients on the base of their medical reports and then referred those patients to us. Then we used different scales to assess primary and secondary outcomes: Diabetes Distress Scale (DDS) and Patient Health Questionnaire (PHQ) to assess primary outcomes, and a Short Health Anxiety Inventory (SHAI), a Revised Version of the Diabetes Quality of Life Questionnaire (DQLQ), and a General Medication Adherence Scale (GMAS) were used to investigate secondary outcomes. Repeated measure ANOVA was used to analyze the results. RESULTS: The findings indicated that patients who received CBT got a significant reduction in their diabetes distress F(1,60) = 222.710, P < 0.001, η2 = .788), depressive symptoms F(1,60) = 94.436, P < 0.001, η2 = .611), health anxiety F(1,60) = 201.915, P < .0.001, η2 = 771), and a significant improvement in their quality of life F(1,60) = 83.352, P < 0.001, η2 = .581), treatment adherence F(1,60) = 67.579, P < 0.001, η2 = .566) and physical activity schedule F(1,60) = 164.245, P < .0.001, η2 = .736 as compared to the patients in waitlist control condition. CONCLUSION: It is concluded that cognitive behavior therapy is an effective and promising intervention for depressive symptoms, diabetes distress, and health anxiety which also helps the person to promote quality of life, treatment adherence and physical activity.

Protective Effects of Choline against Inflammatory Cytokines and Characterization of Transport in Motor Neuron-like Cell Lines (NSC-34).

Choline, a component of the neurotransmitter acetylcholine, is essential for nervous system functions, brain development, and gene expression. In our study, we investigated the protective effect and transport characteristics of choline in amyotrophic lateral sclerosis (ALS) model cell lines. We used the wild-type (WT) motor neuron-like hybrid cell line (NSC-34/hSOD1WT) as a control and the mutant-type (MT; NSC-34/hSOD1G93A) as a disease model. The uptake of [3H]choline was time-, pH-, and concentration-dependent. [3H]Choline transport was sodium-dependent, and, upon pretreatment with valinomycin, induced membrane depolarization. Gene knockdown of Slc44a1 revealed that choline-like transporter 1 (CTL1) mediates the transport of choline. In NSC-34 cell lines, the specific choline transporter inhibitor, hemicholinium-3 demonstrated significant inhibition. Donepezil and nifedipine caused dose-dependent inhibition of [3H]choline uptake by the MT cell line with minimal half inhibitory concentration (IC50) values of 0.14 mM and 3.06 mM, respectively. Four-day pretreatment with nerve growth factor (NGF) resulted in an inhibitory effect on [3H]choline uptake. Choline exerted protective and compensatory effects against cytokines mediators. Hence, the choline transport system CLT1 may act as a potential target for the delivery of novel pharmacological drugs, and the combination of drugs with choline can help treat symptoms related to ALS.

Blood-Brain Barrier Solute Carrier Transporters and Motor Neuron Disease.

Defective solute carrier (SLC) transporters are responsible for neurotransmitter dysregulation, resulting in neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS). We provided the role and kinetic parameters of transporters such as ASCTs, Taut, LAT1, CAT1, MCTs, OCTNs, CHT, and CTL1, which are mainly responsible for the transport of essential nutrients, acidic, and basic drugs in blood-brain barrier (BBB) and motor neuron disease. The affinity for LAT1 was higher in the BBB than in the ALS model cell line, whereas the capacity was higher in the NSC-34 cell lines than in the BBB. Affinity for MCTs was lower in the BBB than in the NSC-34 cell lines. CHT in BBB showed two affinity sites, whereas no expression was observed in ALS cell lines. CTL1 was the main transporter for choline in ALS cell lines. The half maximal inhibitory concentration (IC50) analysis of [3H]choline uptake indicated that choline is sensitive in TR-BBB cells, whereas amiloride is most sensitive in ALS cell lines. Knowledge of the transport systems in the BBB and motor neurons will help to deliver drugs to the brain and develop the therapeutic strategy for treating CNS and neurological diseases.

Antioxidant and Neuroprotective Effects of Paeonol against Oxidative Stress and Altered Carrier-Mediated Transport System on NSC-34 Cell Lines.

Paeonol is a naturally occurring phenolic agent that attenuates neurotoxicity in neurodegenerative diseases. We aimed to investigate the antioxidant and protective effects of paeonol and determine its transport mechanism in wild-type (WT; NSC-34/hSOD1WT) and mutant-type (MT; NSC-34/hSOD1G93A) motor neuron-like amyotrophic lateral sclerosis (ALS) cell lines. Cytotoxicity induced by glutamate, lipopolysaccharides, and H2O2 reduced viability of cell; however, the addition of paeonol improved cell viability against neurotoxicity. The [3H]paeonol uptake was increased in the presence of H2O2 in both cell lines. Paeonol recovered ALS model cell lines by reducing mitochondrial oxidative stress induced by glutamate. The transport of paeonol was time-, concentration-, and pH-dependent in both NSC-34 cell lines. Kinetic parameters showed two transport sites with altered affinity and capacity in the MT cell line compared to the WT cell line. [3H]Paeonol uptake increased in the MT cell line transfected with organic anion transporter1 (Oat1)/Slc22a6 small interfering RNA compared to that in the control. Plasma membrane monoamine transporter (Pmat) was also involved in the uptake of paeonol by ALS model cell lines. Overall, paeonol exhibits neuroprotective activity via a carrier-mediated transport system and may be a beneficial therapy for preventing motor neuronal damage under ALS-like conditions.

Monocarboxylate transporter functions and neuroprotective effects of valproic acid in experimental models of amyotrophic lateral sclerosis.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a devasting neurodegenerative disorder for which no successful therapeutics are available. Valproic acid (VPA), a monocarboxylate derivative, is a known antiepileptic drug and a histone deacetylase inhibitor. METHODS: To investigate whether monocarboxylate transporter 1 (MCT1) and sodium-coupled MCT1 (SMCT1) are altered in ALS cell and mouse models, a cellular uptake study, quantitative real time polymerase chain reaction and western blot parameters were used. Similarly, whether VPA provides a neuroprotective effect in the wild-type (WT; hSOD1WT) and ALS mutant-type (MT; hSOD1G93A) NSC-34 motor neuron-like cell lines was determined through the cell viability assay. RESULTS: [3H]VPA uptake was dependent on time, pH, sodium and concentration, and the uptake rate was significantly lower in the MT cell line than the WT cell line. Interestingly, two VPA transport systems were expressed, and the VPA uptake was modulated by SMCT substrates/inhibitors in both cell lines. Furthermore, MCT1 and SMCT1 expression was significantly lower in motor neurons of ALS (G93A) model mice than in those of WT mice. Notably, VPA ameliorated glutamate- and hydrogen peroxide-induced neurotoxicity in both the WT and MT ALS cell lines. CONCLUSIONS: Together, the current findings demonstrate that VPA exhibits a neuroprotective effect regardless of the dysfunction of an MCT in ALS, which could help develop useful therapeutic strategies for ALS.

Differences of Transport Activity of Arginine and Regulation on Neuronal Nitric Oxide Synthase and Oxidative Stress in Amyotrophic Lateral Sclerosis Model Cell Lines.

L-Arginine, a semi-essential amino acid, was shown to delay dysfunction of motor neurons and to prolong the lifespan, upon analysis of transgenic mouse models of amyotrophic lateral sclerosis (ALS). We investigated the transport function of arginine and neuronal nitric oxide synthase (nNOS) expression after pretreatment with L-arginine in NSC-34 hSOD1WT (wild-type, WT) and hSOD1G93A (mutant-type, MT) cell lines. [3H]L-Arginine uptake was concentration-dependent, voltage-sensitive, and sodium-independent in both cell lines. Among the cationic amino acid transporters family, including system y+, b0,+, B0,+, and y+L, system y+ is mainly involved in [3H]L-arginine transport in ALS cell lines. System b0,+ accounted for 23% of the transport in both cell lines. System B0,+ was found only in MT, and whereas, system y+L was found only in WT. Lysine competitively inhibited [3H]L-arginine uptake in both cell lines. The nNOS mRNA expression was significantly lower in MT than in WT. Pretreatment with arginine elevated nNOS mRNA levels in MT. Oxidizing stressor, H2O2, significantly decreased their uptake; however, pretreatment with arginine restored the transport activity in both cell lines. In conclusion, arginine transport is associated with system y+, and neuroprotection by L-arginine may provide an edge as a possible therapeutic target in the treatment of ALS.

Change in Cationic Amino Acid Transport System and Effect of Lysine Pretreatment on Inflammatory State in Amyotrophic Lateral Sclerosis Cell Model.

Amyotrophic lateral sclerosis (ALS) is a lethal neurological disorder characterized by the deterioration of motor neurons. The aim of this study was to investigate alteration of cationic amino acid transporter (CAT-1) activity in the transport of lysine and the pretreatment effect of lysine on pro-inflammatory states in an amyotrophic lateral sclerosis cell line. The mRNA expression of cationic amino acid transporter 1 was lower in NSC-34/hSOD1G93A (MT) than the control cell line (WT), lysine transport is mediated by CAT-1 in NSC-34 cell lines. The uptake of [3H]L-lysine was Na+-independent, voltage-sensitive, and strongly inhibited by inhibitors and substrates of cationic amino acid transporter 1 (system y+). The transport process involved two saturable processes in both cell lines. In the MT cell line, at a high-affinity site, the affinity was 9.4-fold higher and capacity 24-fold lower than that in the WT; at a low-affinity site, the capacity was 2.3-fold lower than that in the WT cell line. Donepezil and verapamil competitively inhibited [3H]L-lysine uptake in the NSC-34 cell lines. Pretreatment with pro-inflammatory cytokines decreased the uptake of [3H]L-lysine and mRNA expression levels in both cell lines; however, the addition of L-lysine restored the transport activity in the MT cell lines. L-Lysine exhibited neuroprotective effects against pro-inflammatory states in the ALS disease model cell lines. In conclusion, studying the alteration in the expression of transporters and characteristics of lysine transport in ALS can lead to the development of new therapies for neurodegenerative diseases.

Introducing a structured prescription form improves the quality of handwritten prescriptions in limited resource setting of developing countries.

INTRODUCTION: Incomplete or illegible prescriptions can lead to serious errors in administration of the prescribed medication, which can become hazardous. OBJECTIVE: Our aim is to determine if a structured prescription form can improve the quality of handwritten prescription in terms of completeness and legibility. METHODS: We conducted a prospective, non-randomized, time series study of quality of written prescriptions of general practitioners at a tertiary teaching hospital in Peshawar, Pakistan. The study involved an intervention, composed of the introduction of a pre-printed structured prescription form. The data were collected within 4 weeks including a 2-week pre-intervention phase and 2-week post-intervention phase. Completeness, quality of prescriptions and legibility were compared before and after the intervention of the pre-printed structured prescription form. RESULTS: A total of 463 prescriptions were obtained (260 in the pre-intervention phase and 203 in the post-intervention phase). Between pre-intervention phase and post-intervention phase, the Pakistan Medical and Dental Council registration number presence in prescriptions improved from 73.1% to 100% (P < 0.0005). The presence of prescriber's signature improved from 92.7% to 99% (P = 0.001). Drug duration was not missing in 99.5% in post-intervention phase as compared with 90.4% in pre-intervention phase (P < 0.0005). Prescriptions with no legibility problems improved from 76.2% to 94.1% (P < 0.0005). Although not statistically significant, prescriptions in which drug dosage was not missing improved from 85% to 90.6% (P = 0.07). LIMITATIONS: We have a limited single-center study. A larger study in multiple settings is needed to develop adequate evidence for such interventions. Subjective nature of prescription legibility can also be considered as a limitation. CONCLUSION: Structuring a prescription form alone may improve certain aspects of quality of written prescription in terms of completeness and legibility.

Comparatve uric acid lowering studies of allopurinol with an indigenous medicinal plant in rabbits.

The aim of this research was to carry out a comparative study of lowering of uric acid by the use of dried powder of Colchicum luteum and allopathic drug (allopurinol) in rabbits, to determine whether herbal drugs can be used by patients instead of allopathic drugs. The herbal medicine, dried corm powder of Colchicum luteum 2.5 mg/kg/day and dried powder of allopurinol 2 mg/kg/day an allopathic medicine, was used in the study. The results of these medicines were observed in animal model, using 12 adult rabbits, which were divided into three groups A, B and C, respectively, where group C was taken as control. The SPSS version 17 was used for statistical analysis and analysis of variance (ANOVA) was used for comparing the data in different groups and the level of significance was 5%. It was resulted that dried corm of Colchicum luteum significantly reduced the uric acid in adult rabbits as reduced by allopathic medicine--allopurinol. In the light of present research we concluded that the herbal medicines can be used in lieu of allopathic drugs. Thus, the risk of side effects that are associated with the prolonged use of allopathic drugs can be minimized.