In silico, anti-inflammatory and acute toxicological evaluation of an indigenous medicinal plant Pterospermum rubiginosum using Sprague-Dawley rats.

BACKGROUND: Pterospermum rubiginosum has been traditionally used by the tribal inhabitants of Southern India for treating bone fractures and as a local anti-inflammatory agent; however, experimental evidence to support this traditional usage is lacking. The present study aimed to investigate the phytochemical characterization, in silico and in vitro anti-inflammatory evaluation, followed by in vivo toxicological screening of P. rubiginosum methanolic bark extract (PRME). RESULTS: The LCMS evaluation revealed the presence of 80 significant peaks; nearly 50 molecules were identified using the LCMS database. In silico analysis showed notable interactions with inducible nitric oxide synthase (iNOS) and interleukin-6 (IL-6). In vitro gene expression study supported the docking results with significant down-regulation of iNOS, IL-6, and IL-10. PRME was administered orally to the SD rats and was found to be non-toxic up to 1000 mg/kg body weight for 14 days. The antioxidant enzymes catalase and sodium dismutase exhibited an increased value in PRME-administered groups, possibly due to the diverse phytochemical combinations in bark extract. CONCLUSIONS: PRME administration significantly downregulated the gene expression of inflammatory markers, such as iNOS, IL-6, and IL-10. The molecular docking analysis of iNOS and IL-6 supports the in vitro study. In vivo toxicological study of PRME in SD rats was found to be non-toxic up to a concentration of 1000 mg/kg body weight for 14 days.

Engineering resistance against Cucumber mosaic virus in Nicotiana tabacum through virus derived transgene expressing hairpin RNA.

Cucumber mosaic virus (CMV) is the one of notorious virus known for its ubiquitous nature and causes substantial yield loss worldwide. The resistance against the Cucumber mosaic virus (CMV) was envisaged in Nicotiana tabacum transgenic lines by introducing viral gene fragments. The chimeric hairpin RNA constructs incorporating 401 bp of coat protein, 411 bp of replicase protein and 361 bp of 2b gene were developed respectively and transformed into N. tabacum. The regenerated transgenic lines introduced with inverted repeats of CMV gene fragments exhibited enhanced resistance against CMV. The preliminary molecular screening and qPCR confirmed the integration of transgene in the transgenic lines. The spectrum of resistance in transgenic lines was evaluated by challenge inoculation with CMV and the resistance was determined through DAC-ELISA. The complete resistance was achieved in the hpRNA-CP transformant with a very low titre (0.029) of CMV followed by hpRNA-REP (0.099) with no symptoms. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03576-1.

Long-term outcomes of ablation, liver resection, and liver transplant as first-line treatment for solitary HCC of 3 cm or less using an intention-to-treat analysis: A retrospective cohort study.

Background: Curative-intent therapies for hepatocellular carcinoma (HCC) include radiofrequency ablation (RFA), liver resection (LR), and liver transplantation (LT). Controversy exists in treatment selection for early-stage tumours. We sought to evaluate the oncologic outcomes of patients who received either RFA, LR, or LT as first-line treatment for solitary HCC ≤ 3 cm in an intention-to-treat analysis. Materials and methods: All patients with solitary HCC ≤ 3 cm who underwent RFA, LR, or were listed for LT between Feb-2000 and Nov-2018 were analyzed. Cox regression analysis was then performed to compare intention-to-treat (ITT) survival by initial treatment allocation and disease-free survival (DFS) by treatment received in patients eligible for all three treatments. Results: A total of 119 patients were identified (RFA n = 83; LR n = 25; LT n = 11). The overall intention-to-treat survival was similar between the three groups. The overall DFS was highest for the LT group. This was significantly higher than RFA (p = 0.02), but not statistically significantly different from LR (p = 0.14). After multivariable adjustment, ITT survival was similar in the LR and LT groups relative to RFA (LR HR:1.13, 95%CI 0.33-3.82; p = 0.80; LT HR:1.39, 95%CI 0.35-5.44; p = 0.60). On multivariable DFS analysis, only LT was better relative to RFA (LR HR:0.52, 95%CI 0.26-1.02; p = 0.06; LT HR:0.15, 95%CI 0.03-0.67; p = 0.01). Compared to LR, LT was associated with a numerically lower hazard on multivariable DFS analysis, though this did not reach statistical significance (HR 0.30, 95%CI 0.06-1.43; p = 0.13). Conclusion: For treatment-naïve patients with solitary HCC ≤ 3 cm who are eligible for RFA, LR, and LT, adjusted ITT survival is equivalent amongst the treatment modalities, however, DFS is better with LR and LT, compared with RFA. Differences in recurrence between treatment modalities and equipoise in ITT survival provides support for a future prospective trial in this setting.

Co-existence of chlorosis inducing strain of Cucumber mosaic virus with tospoviruses on hot pepper (Capsicum annuum) in India.

Cucumo- and tospoviruses are the most destructive viruses infecting hot pepper (chilli). A diagnostic survey was conducted to assess the prevalence of cucumo and tospoviruses in chilli growing tracts of Tamil Nadu. Infected plants showing mosaic with chlorotic and necrotic rings, veinal necrosis, mosaic mottling, leaf filiformity and malformation were collected. Molecular indexing carried out through reverse transcription polymerase chain reaction (RT-PCR) with coat protein gene specific primer of Cucumber mosaic virus (CMV) and tospovirus degenerate primer corresponding to the L segment (RdRp). Ostensibly, amplifications were observed for both CMV and tospoviruses as sole as well for mixed infections. The sequence analysis indicated that the Capsicum chlorosis virus (CaCV) and Groundnut bud necrosis virus (GBNV) to be involved with CMV in causing combined infections. The co-infection of CMV with CaCV was detected in 10.41% of the symptomatic plant samples and combined infection of CMV with GBNV was recorded in around 6.25% of the symptomatic plants surveyed. The amino acid substitution of Ser129 over conserved Pro129 in coat protein of CMV implies that CMV strain involved in mixed infection as chlorosis inducing strain. Further, the electron microscopy of symptomatic plant samples explicated the presence of isometric particles of CMV and quasi spherical particles of tospoviruses. This is the first molecular evidence for the natural co-existence of chlorosis inducing CMV strain with CaCV and GBNV on hot pepper in India.

Characterization of cucumber mosaic virus (CMV) subgroup IB infecting chilli in Tamil Nadu, India.

CMV (cucumber mosaic virus) is the most primitive virus infecting chilli (Capsicum annuum. L). The mosaic incidence with leaf filiformity, mosaic mottling and stunted growth was observed in major chilli growing regions of Tamil Nadu. CMV sap was inoculated on chilli, cowpea, bitter gourd, bottle gourd, ridge gourd, banana, cucumber, Nicotiana and Chenopodium plants. Host range studies revealed that CMV produced localized infection on Nicotiana and systemic symptoms on most of the test plants. The occurrence of CMV was confirmed through DAC-ELISA and RT-PCR analysis. Host plant samples tested with DAC-ELISA showed strong reaction with 1.7 optical density. For molecular characterization, total RNA isolated from infected plants used in RT-PCR with CMV specific primers. The specific amplicons were cloned and sequenced. The complete genome sequencing depicts CMV-RNA1 consist of 3339 nucleotides (nt), RNA2 and RNA3 with 3052nt and 2027nt respectively. Phylogenetic and nucleotide sequence analysis showed TN CMV isolates closely associated with subgroup IB rather than subgroup IA and II. Comparative sequence analysis indicates replicase protein to be more variable among five genes. CP sequence analysis showed 97-98 per cent identity with subgroup IB strains, 92-93 per cent identity with subgroup IA strains and 81-82 per cent identity with subgroup II strains. CMV-RNA3 was predicted to have recombination with Indian black pepper isolate (KU947031) between 165-505nt and Egyptian tomato isolate (KX014666) between 165-506nt positions.

Theoretical Analysis of Voltammetry at a Rotating Disk Electrode in the Absence of Supporting Electrolyte.

The mathematical model of Oldham J. Phys. Chem. B 2000 , 104 , 4703 for rotating disc electrode in an unsupported system is discussed. This article presents a new analytical method for the calculation of concentration at a rotating disc electrode controlled by diffusion, convection, and migration. This model contains a steady-state nonlinear differential equation in a three-ion system under the assumption that all the ions have the same diffusivity. The homotopy perturbation method is employed to solve the nonlinear governing equation, where the ionic concentration and current are obtained analytically, in terms of charge numbers. The comparison between the analytical results of this study and previous studies confirms that the result of the proposed method is in stronger agreement with numerical simulations than other analytical methods.

Nested and TaqMan® probe based quantitative PCR for the diagnosis of Ca. Phytoplasma in coconut palms.

The root (wilt) disease caused by phytoplasma (Ca. Phytoplasma) is one of the major and destructive occurs in coconut gardens of Southern India. As this organism could not be cultured in vitro, the early detection in the palm is very much challenging. Hence, proper early diagnosis and inoculum assessment relay mostly on the molecular techniques namely nested and quantitative PCR (qPCR). So, the present study qPCR assay conjugated with TaqMan® probe was developed which is a rapid, sensitive method to detect the phytoplasma. For the study, samples from different parts of infected coconut palms viz., spindle leaflets, roots and the insect vector-leaf hopper (Proutista moesta) were collected and assessed by targeting 16S rRNA gene. Further, nested PCR has been carried out using p1/p7 and fU5/rU3 primers and resulted in the amplification product size of 890 bp. From this amplified product, specifically a target of 69 bp from the 16S rRNA gene region has been detected through primers conjugated with Taqman probe in a step one instrument. The results indicated that the concentration of phytoplasma was more in spindle leaflets (8.9 × 105 g of tissue) followed by roots (7.4 × 105 g of tissue). Thus, a qPCR approach for detection and quantification of coconut phytoplasma was more advantageous than other PCR methods in terms of sensitivity and also reduced risk of cross contamination in the samples. Early diagnosis and quantification will pave way for the healthy coconut saplings selection and management under field conditions.

Production of polyclonal antibodies for Capsicum chlorosis virus (CaCV) infecting chilli in India through recombinant nucleocapsid protein expression and its application.

Bud necrosis and chlorotic spots causing virus affecting chilli crop in Tamil Nadu (India) was identified as Capsicum chlorosis virus (CaCV). Specific primers were used for amplification and sequencing of the nucleocapsid protein (NP) gene. Polyclonal antibody against the bacterially expressed NP from the CaCV-TN-CBE isolate was produced using recombinant DNA technology. NP gene was subcloned into the pET-28a (+) vector and expressed by transformation in BL21 (DE3) pLysS. The expressed protein was about ∼34 kDa and was confirmed through western blot analysis using Groundnut bud necrosis virus (GBNV) polyclonal antiserum from ICRISAT, India. The purified recombinant protein was used to immunize rabbits to generate CaCV-specific polyclonal antiserum. The sensitivity levels of polyclonal antiserum thus raised was assayed through indirect ELISA or direct antigen coating (DAC)-ELISA using the recombinant protein as antigen. The recombinant antiserum produced in this study successfully detected the natural infection of CaCV on chilli plants collected from the field as well as on cowpea plants artificially inoculated with CaCV by using DAC-ELISA, DIBA and western blotting.

Unprecedented homotopy perturbation method for solving nonlinear equations in the enzymatic reaction of glucose in a spherical matrix.

The theory of glucose-responsive composite membranes for the planar diffusion and reaction process is extended to a microsphere membrane. The theoretical model of glucose oxidation and hydrogen peroxide production in the chitosan-aliginate microsphere has been discussed in this manuscript for the first time. We have successfully reported an analytical derived methodology utilizing homotopy perturbation to perform the numerical simulation. The influence and sensitive analysis of various parameters on the concentrations of gluconic acid and hydrogen peroxide are also discussed. The theoretical results enable to predict and optimize the performance of enzyme kinetics.

Part-2: Analytical Expressions of Concentrations of Glucose, Oxygen, and Gluconic Acid in a Composite Membrane for Closed-Loop Insulin Delivery for the Non-steady State Conditions.

A mathematical model developed by Abdekhodaie and Wu (J Membr Sci 335:21-31, 2009), which describes a dynamic process involving an enzymatic reaction and diffusion of reactants and product inside glucose-sensitive composite membrane has been discussed. This theoretical model depicts a system of non-linear non-steady state reaction diffusion equations. These equations have been solved using new approach of homotopy perturbation method and analytical solutions pertaining to the concentrations of glucose, oxygen, and gluconic acid are derived. These analytical results are compared with the numerical results, and limiting case results for steady state conditions and a good agreement is observed. The influence of various kinetic parameters involved in the model has been presented graphically. Theoretical evaluation of the kinetic parameters like the maximal reaction velocity (V max) and Michaelis-Menten constants for glucose and oxygen (K g and K ox) is also reported. This predicted model is very much useful for designing the glucose-responsive composite membranes for closed-loop insulin delivery.

Calcium flux-independent NMDA receptor activity is required for Aß oligomer-induced synaptic loss.

Synaptic loss is one of the major features of Alzheimer's disease (AD) and correlates with the degree of dementia. N-methyl-D-aspartate receptors (NMDARs) have been shown to mediate downstream effects of the ß-amyloid peptide (Aß) in AD models. NMDARs can trigger intracellular cascades via Ca(2+) entry, however, also Ca(2+)-independent (metabotropic) functions of NMDARs have been described. We aimed to determine whether ionotropic or metabotropic NMDAR signaling is required for the induction of synaptic loss by Aß. We show that endogenous Aß as well as exogenously added synthetic Aß oligomers induced dendritic spine loss and reductions in pre- and postsynaptic protein levels in hippocampal slice cultures. Synaptic alterations were mitigated by blocking glutamate binding to NMDARs using NMDAR antagonist APV, but not by preventing ion flux with Ca(2+) chelator BAPTA or open-channel blockers MK-801 or memantine. Aß increased the activity of p38 MAPK, a kinase involved in long-term depression and inhibition of p38 MAPK abolished the loss of dendritic spines. Aß-induced increase of p38 MAPK activity was prevented by APV but not by BAPTA, MK-801 or memantine treatment highlighting the role of glutamate binding to NMDARs but not Ca(2+) flux for synaptic degeneration by Aß. We further show that treatment with the G protein inhibitor pertussis toxin (PTX) did not prevent dendritic spine loss in the presence of Aß oligomers. Our data suggest that Aß induces the activation of p38 MAPK and subsequent synaptic loss through Ca(2+) flux- and G protein-independent mechanisms.

Analysis of mathematical modelling on potentiometric biosensors.

A mathematical model of potentiometric enzyme electrodes for a nonsteady condition has been developed. The model is based on the system of two coupled nonlinear time-dependent reaction diffusion equations for Michaelis-Menten formalism that describes the concentrations of substrate and product within the enzymatic layer. Analytical expressions for the concentration of substrate and product and the corresponding flux response have been derived for all values of parameters using the new homotopy perturbation method. Furthermore, the complex inversion formula is employed in this work to solve the boundary value problem. The analytical solutions obtained allow a full description of the response curves for only two kinetic parameters (unsaturation/saturation parameter and reaction/diffusion parameter). Theoretical descriptions are given for the two limiting cases (zero and first order kinetics) and relatively simple approaches for general cases are presented. All the analytical results are compared with simulation results using Scilab/Matlab program. The numerical results agree with the appropriate theories.

Analytical expressions for the steady-state concentrations of glucose, oxygen and gluconic acid in a composite membrane for closed-loop insulin delivery.

The mathematical model of Abdekhodaie and Wu (J Membr Sci 335:21-31, 2009) of glucose-responsive composite membranes for closed-loop insulin delivery is discussed. The glucose composite membrane contains nanoparticles of an anionic polymer, glucose oxidase and catalase embedded in a hydrophobic polymer. The model involves the system of nonlinear steady-state reaction-diffusion equations. Analytical expressions for the concentration of glucose, oxygen and gluconic acid are derived from these equations using the Adomian decomposition method. A comparison of the analytical approximation and numerical simulation is also presented. An agreement between analytical expressions and numerical results is observed.

Active vaccination with ankyrin G reduces ß-amyloid pathology in APP transgenic mice.

Serum antibodies against amyloid-ß peptide (Aß) in humans with or without diagnosis of Alzheimer's disease (AD) indicate the possibility of immune responses against brain antigens. In an unbiased screening for antibodies directed against brain proteins, we found in AD patients high serum levels of antibodies against the neuronal cytoskeletal protein ankyrin G (ankG); these correlated with slower rates of cognitive decline. Neuronal expression of ankG was higher in AD brains than in nondemented age-matched healthy control subjects. AnkG was present in exosomal vesicles, and it accumulated in ß-amyloid plaques. Active immunization with ankG of arcAß transgenic mice reduced brain ß-amyloid pathology and increased brain levels of soluble Aß(42). AnkG immunization induced a reduction in ß-amyloid pathology, also in Swedish transgenic mice(.) Anti-ankG monoclonal antibodies reduced Aß-induced loss of dendritic spines in hippocampal ArcAß organotypic cultures. Together, these data established a role for ankG in the human adaptive immune response against resident brain proteins, and they show that ankG immunization reduces brain ß-amyloid and its related neuropathology.

Effect of pyridoxine supplementation on recurrent stone formers.

Twelve recurrent stone formers with hyperoxaluria were administered pyridoxine-HCl (10 mg/day) daily for a period of 180 days. The pyridoxine status of the patients, as assessed by their erythrocyte transaminase activation indexes, improved significantly (p less than 0.001) after 180 days of supplementation as compared with the basal levels. Although urinary oxalate decreased significantly (p less than 0.05) by the 90th day of pyridoxine therapy, other parameters, e.g., urinary calcium, phosphorus, and creatinine, remained unaltered. Significant correlation was observed between erythrocyte glutamate pyruvate transaminase (EGPT) or erythrocyte glutamate oxaloacetate transaminase (EGOT) activation index and urinary oxalate excretion (p less than 0.01). Pyridoxine in low doses (10 mg/day) is of therapeutic value for hyperoxaluric stone formers.

Operative management problems in nephrobronchial fistula.

During left subcapsular nephrectomy in a patient with nephrobronchial fistula, the purulent material entered the bronchial tree through the fistulous tracts and flooded the dependent right lung. Patchy atelectasis and later massive consolidation of the lower lobe of the right lung ensued ultimately causing her death. A plea is made to use double-lumen endobronchial tubes for anesthesia to prevent such spillover of purulent secretions to the contralateral lung from the kidney. The fistulous tracts should be divided as the first step during surgery, before mobilization of the kidney. In a suspected case, retrograde pyelography should be done preferably under local or regional anesthesia, so that the patient could cough out the contrast material and purulent secretion that might enter the bronchial tree under pressure during the procedure.

Peripelvic extravasation and formation of perinephric urinoma after cystoscopy.

A hitherto undescribed complication of cystoscopy is reported wherein peripelvic extravasation and perinephric collection of urine resulted from conventional cystoscopic examination in a patient with a small-capacity bladder and vesicoureteral reflux. Prompt recognition, institution of indwelling catheter drainage, and massive antibiotic therapy led to spontaneous resolution of the flank mass. In similar cases, we suggest the use of an Iglesias resectoscope with a continuous flow and suction evacuation, even for cystoscopy and bladder biopsy.