Specific consistency score for rational selection of epilepsy resection surgery candidates.

OBJECTIVE: Degree of indication for epilepsy surgery is determined by taking multiple factors into account. This study aimed to investigate the usefulness of the Specific Consistency Score (SCS), a proposed score for focal epilepsy to rate the indication for epilepsy focal resection. METHODS: This retrospective cohort study included patients considered for resective epilepsy surgery in Kyoto University Hospital from 2011 to 2022. Plausible epileptic focus was tentatively defined. Cardinal findings were scored based on specificity and consistency with the estimated laterality and lobe. The total points represented SCS. The association between SCS and the following clinical parameters was assessed by univariate and multivariate analysis: (1) probability of undergoing resective epilepsy surgery, (2) good postoperative seizure outcome (Engel I and II or Engel I only), and (3) lobar concordance between the noninvasively estimated focus and intracranial electroencephalographic (EEG) recordings. RESULTS: A total of 131 patients were evaluated. Univariate analysis revealed higher SCS in the (1) epilepsy surgery group (8.4 [95% confidence interval (CI) = 7.8-8.9] vs. 4.9 [95% CI = 4.3-5.5] points; p < .001), (2) good postoperative seizure outcome group (Engel I and II; 8.7 [95% CI = 8.2-9.3] vs. 6.4 [95% CI = 4.5-8.3] points; p = .008), and (3) patients whose focus defined by intracranial EEG matched the noninvasively estimated focus (8.3 [95% CI = 7.3-9.2] vs. 5.4 [95% CI = 3.5-7.3] points; p = .004). Multivariate analysis revealed areas under the curve of .843, .825, and .881 for Parameters 1, 2, and 3, respectively. SIGNIFICANCE: SCS provides a reliable index of good indication for resective epilepsy surgery and can be easily available in many institutions not necessarily specializing in epilepsy.

Gene therapy for alopecia in type II rickets model rats using vitamin D receptor-expressing adenovirus vector.

Type II rickets is a hereditary disease caused by a mutation in the vitamin D receptor (VDR) gene. The main symptoms of this disease are bone dysplasia and alopecia. Bone dysplasia can be ameliorated by high calcium intake; however, there is no suitable treatment for alopecia. In this study, we verified whether gene therapy using an adenoviral vector (AdV) had a therapeutic effect on alopecia in Vdr-KO rats. The VDR-expressing AdV was injected into six 7-week-old female Vdr-KO rats (VDR-AdV rats). On the other hand, control-AdV was injected into 7-week-old female rats (control-AdV rats); non-infected Vdr-KO rats (control rats) were also examined. The hair on the backs of the rats was shaved with hair clippers, and VDR-AdV or control-AdV was intradermally injected. Part of the back skin was collected from each rat after AdV administration. Hair follicles were observed using hematoxylin and eosin staining, and VDR expression was examined using immunostaining and western blotting. VDR-AdV rats showed significant VDR expression in the skin, enhanced hair growth, and low cyst formation, whereas control-AdV and non-infected rats did not show any of these effects. The effect of VDR-AdV lasted for nearly 60 days. These results indicate that gene therapy using VDR-AdV may be useful to treat alopecia associated with type II rickets, if multiple injections are possible after a sufficient period of time.

Effect of Nonsupplemented Low-Protein Diet on the Initiation of Renal Replacement Therapy in Stage 4 and 5 Chronic Kidney Disease: A Retrospective Multicenter Cohort Study in Japan.

OBJECTIVE: In subjects with chronic kidney disease (CKD), the effect of low-protein diet (LPD) is expected to alleviate uremic symptoms. However, whether LPD is effective in preventing loss of kidney function is controversial. The aim of this study was to evaluate the association between LPD and renal outcomes. METHODS: We conducted a multicenter cohort study of 325 patients who suffered CKD stage 4 and 5 with eGFR ≥10 mL/min/1.73 m,2 between January 2008 and December 2014. The primary diseases of the patients were chronic glomerulonephritis (47.7%), nephrosclerosis (16.9%), diabetic nephropathy (26.2%), and others (9.2%). The patients were divided into four groups, based on the mean protein intake (PI)/day, group 1 (n = 76): PI < 0.5 g/kg ideal body weight/day, group 2 (n = 56): 0.5 ≤ PI < 0.6 g/kg/day, group 3 (n = 110): 0.6 ≤ PI < 0.8 g/kg/day, group 4 (n = 83): PI ≥ 0.8 g/kg/day. Dietary supplementation with essential amino acids and ketoanalogues was not used. The outcome measure was occurrence of renal replacement therapy (RRT) (hemodialysis, peritoneal dialysis, renal transplantation (excluding preemptive transplantation)) and all-cause mortality until December 2018. Cox regression models were used to examine whether LPD was associated with the risk of outcomes. RESULTS: During a mean follow-up of 4.1 ± 2.2 years. Thirty-three patients (10.2%) died of all causes, 163 patients (50.2%) needed to start RRT, and 6 patients (1.8%) received a renal transplant. LPD therapy of 0.5 g/kg/day or less was significantly related to a lower risk of RRT and all-cause mortality [Hazard ratio = 0.656; 95% confidence interval, 0.438 to 0.984, P = .042]. CONCLUSIONS: These results suggest that non-supplemented LPD therapy of 0.5 g/kg/day or less may prolong the initiation of RRT in stage 4 and 5 CKD patients.

Expression of Rat Cyp27b1 in HepG2 Cells Using Adenovirus Vector and Its Application to Evaluation of Self-Made and Commercially Available Anti-Cyp27b1 Antibodies.

Rat Cyp27b1 was successfully expressed in HepG2 cells using an adenovirus vector. High vitamin D 1α-hydroxylation activity was detected in them, whereas no activity was observed in non-infected cells. Similarly, vitamin D 1α-hydroxylation activity was also observed in HepG2 cells expressing Cyp27b1-Flag, which is tagged with a Flag at the C-terminus of Cyp27b1. Western blot analysis using an anti-Flag antibody showed a clear band of Cyp27b1-Flag. Next, we screened three types of anti-Cyp27b1 antibodies, which consist of two commercially available antibodies and our self-made antibody using Cyp27b1- or Cyp27b1-Flag expressing HepG2 cell lysate as a positive control. Surprisingly, Western blot analysis revealed that two commercially available antibodies did not detect Cyp27b1 but specifically detect other proteins. In contrast, our self-made antisera specifically detected Cyp27b1 and Cyp27b1-Flag in the HepG2 cells expressing Cyp27b1 or Cyp27b1-Flag. These commercially available antibodies have been used for the detection of Cyp27b1 by Western blotting and immunohistochemistry. Our results suggest that those data should be reanalyzed.

Functional analysis of vitamin D receptor (VDR) using adenovirus vector.

Recently, we generated type II rickets model rats, including Vdr(R270L), Vdr(H301Q), Vdr(R270L/H301Q), and Vdr-knockout (KO), by genome editing. All generated animals showed symptoms of rickets, including growth retardation and abnormal bone formation. Among these, only Vdr-KO rats exhibited abnormal skin formation and alopecia. To elucidate the relationship between VDR function and rickets symptoms, each VDR was expressed in human HaCaT-VDR-KO cells using an adenovirus vector. We also constructed an adenovirus vector expressing VDR(V342M) corresponding to human VDR(V346M) which causes alopecia. We compared the nuclear translocation of VDRs after adding 1α,25-dihydroxyvitamin D3 (1,25D3) or 25-hydroxyvitamin D3 (25D3) at final concentrations of 10 and 100 nM, respectively. Both 25D3 and 1,25D3 induced the nuclear translocation of wild type VDR and VDR(V342M). Conversely, VDR(R270L) translocation was observed in the presence of 100 nM 25D3, with almost no translocation following treatment with 10 nM 1,25D3. VDR(R270L/H301Q) failed to undergo nuclear translocation. These results were consistent with their affinity for each ligand. Notably, VDR(R270L/H301Q) may exist in an unliganded form under physiological conditions, and factors interacting with VDR(R270L/H301Q) may be involved in the hair growth cycle. Thus, this novel system using an adenovirus vector could be valuable in elucidating vitamin D receptor functions.

CREG1 improves diet-induced obesity via uncoupling protein 1-dependent manner in mice.

Thermogenic brown and beige adipocytes express uncoupling protein 1 (UCP1) and stimulate energy metabolism, protecting against obesity and metabolic diseases such as type 2 diabetes and hyperlipidemia. Cellular repressor of E1A-stimulated genes 1 (CREG1) can stimulate thermogenic fat formation, induce UCP1, and reduce diet-induced obesity (DIO) in mice at normal room temperature. In this study, we investigated the effect of CREG1 administration and the importance of UCP1 in DIO inhibition under thermoneutral conditions at 30°C, which attenuate thermogenic fat formation. Interestingly, subcutaneous administration of recombinant CREG1 protein via an osmotic pump in C57BL/6J mice for four weeks increased UCP1 expression in interscapular brown adipose tissue (IBAT), inhibited visceral white fat hypertrophy with partial browning, and reduced DIO compared to that in PBS-treated mice. The mRNA expression of energy metabolism-related genes was significantly increased in the IBAT of CREG1-treated mice compared to that in PBS-treated mice. In contrast, adipocyte-specific overexpression of CREG1 failed to improve DIO in UCP1-knockout mice at thermoneutrality. Our results indicate the therapeutic potential of CREG1 administration for obesity under thermogenic fat-attenuating conditions and highlight the indispensable role of UCP1 in the DIO-inhibitory effect of CREG1.

CREG1 administration stimulates BAT thermogenesis and improves diet-induced obesity in mice.

Brown and beige adipocytes, which express thermogenic uncoupling protein-1 (UCP1), stimulate glucose and lipid metabolism, improving obesity and metabolic diseases such as type 2 diabetes and hyperlipidemia. Overexpression of cellular repressor of E1A-stimulated genes 1 (CREG1) promotes adipose tissue browning and inhibits diet-induced obesity (DIO) in mice. In this study, we investigated the effects of CREG1 administration on DIO inhibition and adipose browning. Subcutaneous administration of recombinant CREG1 protein to C57BL/6 mice stimulated UCP1 expression in interscapular brown adipose tissue (IBAT) and improved DIO, glucose tolerance and fatty liver compared with those in phosphate-buffered saline-treated mice. Injection of Creg1-expressing adenovirus into inguinal white adipose tissue (IWAT) significantly increased browning and mRNA expression of beige adipocyte marker genes compared with that in mice injected with control virus. The effect of Creg1 induction on beige adipocyte differentiation was supported in primary culture using preadipocytes isolated from IWAT of Creg1-transgenic mice compared with that of wild-type mice. Our results indicate a therapeutic effect of CREG1 on obesity and its associated pathology and a potential of CREG1 to stimulate brown/beige adipocyte formation.

Effect of tolvaptan in Japanese patients with autosomal dominant polycystic kidney disease: a post hoc analysis of TEMPO 3:4 and TEMPO Extension Japan.

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is a progressive condition that eventually leads to end-stage renal disease. A phase 3 trial of tolvaptan (TEMPO 3:4; NCT00428948) and its open-label extension (TEMPO Extension Japan: TEMPO-EXTJ; NCT01280721) were conducted in patients with ADPKD. In this post hoc analysis, effects on renal function and the safety profile of tolvaptan were assessed over a long-term period that included the 3-year TEMPO 3:4 and the approximately 3-year TEMPO-EXTJ trials. METHODS: Patients from Japanese trial sites who completed TEMPO 3:4 were offered participation in TEMPO-EXTJ. Patients whose efficacy parameters were measured at year 2 in TEMPO-EXTJ for efficacy evaluation were included. The annual slope of the estimated glomerular filtration rate (eGFR) and growth in total kidney volume (TKV) were analyzed. RESULTS: In patients who received tolvaptan in TEMPO 3:4 and TEMPO-EXTJ, the annual slope of eGFR (mL/min/1.73 m2) was - 3.480 in TEMPO 3:4 and - 3.417 in TEMPO-EXTJ, with no apparent effect of an approximately 3.6-month off-treatment interval between the two trials. In patients who received a placebo in TEMPO 3:4 before initiating tolvaptan in TEMPO-EXTJ, the slope of eGFR was significantly less steep from TEMPO 3:4 (- 4.287) to TEMPO-EXTJ (- 3.364), a difference of 0.923 (P = 0.0441). CONCLUSION: The TEMPO-EXTJ trial supports a sustained beneficial effect of tolvaptan on eGFR. In patients who received a placebo in TEMPO 3:4, initiation of tolvaptan in TEMPO-EXTJ was associated with a significant slowing of eGFR decline.

Preservation of kidney function irrelevant of total kidney volume growth rate with tolvaptan treatment in patients with autosomal dominant polycystic kidney disease.

BACKGROUND: Tolvaptan slowed the rates of total kidney volume (TKV) growth and renal function decline over a 3-year period in patients with autosomal dominant polycystic kidney disease (ADPKD) enrolled in the Tolvaptan Efficacy and Safety in Management of Autosomal Dominant Polycystic Kidney Disease and Its Outcomes (TEMPO) 3:4 trial (NCT00428948). In this post hoc analysis of Japanese patients from TEMPO 3:4, we evaluated whether the effects of tolvaptan on TKV and on renal function are interrelated. METHODS: One hundred and forty-seven Japanese patients from TEMPO 3:4 were included in this analysis (placebo, n = 55; tolvaptan, n = 92). Tolvaptan-treated patients were stratified into the responder group (n = 37), defined as tolvaptan-treated patients with a net decrease in TKV from baseline to year 3, and the non-responder group (n = 55), defined as tolvaptan-treated patients with a net increase in TKV. RESULTS: Mean changes during follow-up in the placebo, responder, and non-responder groups were 16.99%, - 8.33%, and 13.95%, respectively, for TKV and - 12.61, - 8.47, and - 8.58 mL/min/1.73 m2, respectively, for estimated glomerular filtration rate (eGFR). Compared with the placebo group, eGFR decline was significantly slowed in both the responder and non-responder groups (P < 0.05). CONCLUSION: Tolvaptan was effective in slowing eGFR decline, regardless of TKV response, over 3 years in patients with ADPKD in Japan. Treatment with tolvaptan may have beneficial effects on slowing of renal function decline even in patients who have not experienced a reduction in the rate of TKV growth by treatment with tolvaptan.

Efficacy and safety of oral tolvaptan in patients undergoing hemodialysis: a Phase 2, double-blind, randomized, placebo-controlled trial.

BACKGROUND: Loop diuretics are used to manage fluid retention in patients with end-stage kidney disease undergoing hemodialysis (HD). This randomized, double-blind, placebo-controlled, Phase 2 trial evaluated the efficacy and safety of tolvaptan, a vasopressin V2 receptor antagonist, in Japanese HD patients. METHODS: A total of 124 patients (24-h urine volume ≥500 mL) on thrice-weekly HD were randomized to receive oral tolvaptan 15 mg/day (n = 40), tolvaptan 30 mg/day (n = 40) or placebo (n = 44) for 24 weeks. Efficacy endpoints were change from baseline in 24-h urine volume, total fluid removal by HD per week and interdialytic weight gain (IDWG). Safety was assessed via the incidence of treatment-emergent adverse events (TEAEs). RESULTS: At treatment end, the difference (95% confidence interval) from the placebo group in the mean change from baseline in 24-h urine volume was significant in the tolvaptan 15 mg {429.1 mL [95% confidence interval (CI) 231.0, 627.2]; P < 0.0001} and 30 mg [371.6 mL (95% CI 144.1, 599.2); P = 0.0017] groups. The mean changes from baseline in total fluid removal by HD and IDWG were not significantly different in the tolvaptan groups versus the placebo group. Although the proportion of patients with TEAEs was lower in the placebo group (77.3%) than in the tolvaptan groups (92.3%), tolvaptan was safe and well-tolerated during the study period. CONCLUSIONS: Tolvaptan significantly sustained diuretic action for 24 weeks in HD patients but did not reduce total fluid removal by HD per week and IDWG to the same extent.

Impaired adaptation of energy intake induces severe obesity in aged mice on a high-fat diet.

High-fat diet (HFD) feeding induces inflammation in various tissues, including the nodose ganglion and hypothalamus, resulting in obesity and metabolic disorders. In this study, we investigated the effect of short-term HFD on aged and young mice. Aged mice easily gained weight during short-term HFD feeding, and required many days to adapt their energy intake. One-day HFD in aged mice induced inflammation in the distal colon, but not in the nodose ganglion or hypothalamus. The anorexic effect of glucagon-like peptide-1 (GLP-1) was attenuated in aged mice. Intraperitoneal administration of GLP-1 did not induce expression of genes that regulate feeding in the hypothalamus of aged mice. mRNA expression of the gene encoding the GLP-1 receptor (Glp1r) in the nodose ganglion was significantly lower in aged mice than in young mice. Our findings suggest that adaptation of energy intake regulation was attenuated in aged mice, causing them to become obese in response to short-term HFD feeding.

Repurposing existing drugs: identification of irreversible IMPDH inhibitors by high-throughput screening.

Inosine 5'-monophosphate dehydrogenase (IMPDH) is an essential enzyme for the production of guanine nucleotides. Disruption of IMPDH activity has been explored as a therapeutic strategy for numerous purposes, such as for anticancer, immunosuppression, antiviral, and antimicrobial therapy. In the present study, we established a luciferase-based high-throughput screening system to identify IMPDH inhibitors from our chemical library of known bioactive small molecules. The screening of 1400 compounds resulted in the discovery of three irreversible inhibitors: disulfiram, bronopol, and ebselen. Each compound has a distinct chemical moiety that differs from other reported IMPDH inhibitors. Further evaluation revealed that these compounds are potent inhibitors of IMPDHs with kon values of 0.7 × 104 to 9.3 × 104 M-1·s-1. Both disulfiram and bronopol exerted similar degree of inhibition to protozoan and mammalian IMPDHs. Ebselen showed an intriguing difference in mode of inhibition for different IMPDHs, with reversible and irreversible inhibition to each Cryptosporidium parvum IMPDH and human IMPDH type II, respectively. In the preliminary efficacy experiment against cryptosporidiosis in severe combined immunodeficiency (SCID) mouse, a decrease in the number of oocyst shed was observed upon the oral administration of disulfiram and bronopol, providing an early clinical proof-of-concept for further utilization of these compounds as IMPDH inhibitors.

Analysis of peripheral ghrelin signaling via the vagus nerve in ghrelin receptor-restored GHSR-null mice.

The vagus nerve connects peripheral organs to the central nervous system (CNS), and gastrointestinal hormones transmit their signals to the CNS via the vagal afferent nerve. Ghrelin, a gastric-derived orexigenic peptide, stimulates food intake by transmitting starvation signals via the vagus nerve. To understand peripheral ghrelin signaling via the vagus nerve, we investigated the ghrelin receptor (GHSR)-null mouse. For this purpose, we tried to produce mice in which GHSR was selectively expressed in the hindbrain and vagus nerve. GHSR was expressed in some nodose ganglion neurons in these mice, but GHSR-expressing neurons were less abundant than in wild-type mice. Intraperitoneal administration of ghrelin did not induce food intake or growth hormone release, but did increase blood glucose levels. Our findings suggest that the abundance of GHSR-expressing neurons in the nodose ganglion is critical for peripheral administration of ghrelin-induced food intake and growth hormone release via the vagus nerve.

Restoration of metabolic inflammation-related ghrelin resistance by weight loss.

High-fat diet (HFD)-induced metabolic inflammation in the central and peripheral organs contributes to the pathogenesis of obesity. Long-term HFD blunts signaling by ghrelin, a gastric-derived orexigenic peptide, in the vagal afferent nerve via a mechanism involving in situ activation of inflammation. This study was undertaken to investigate whether ghrelin resistance is associated with progressive development of metabolic inflammation. In mice, ghrelin's orexigenic activity was abolished 2-4 weeks after the commencement of HFD (60% of energy from fat), consistent with the timing of accumulation and activation of macrophages and microglia in the nodose ganglion and hypothalamus. Calorie-restricted weight loss after 12-week HFD feeding restored ghrelin responsiveness and alleviated the upregulation of macrophage/microglia activation markers and inflammatory cytokines. HSP72, a chaperone protein, was upregulated in the hypothalamus of HFD-fed mice, potentially contributing to prevention of irreversible neuron damage. These results demonstrate that ghrelin resistance is reversible following reversal of the HFD-induced inflammation and obesity phenotypes.

Long-term safety profile of tolvaptan in autosomal dominant polycystic kidney disease patients: TEMPO Extension Japan Trial.

AIM: The aim of this trial (ClinicalTrials.gov identifier: NCT01280721) was to investigate the long-term safety profile of tolvaptan in Japanese patients with autosomal dominant polycystic kidney disease (ADPKD). METHODS: This open-label multicenter trial was conducted to examine adverse drug reactions (ADRs) related to tolvaptan up to an additional 3 years in 135 Japanese patients who participated in the Tolvaptan Efficacy and Safety in Management of Autosomal Dominant Polycystic Kidney Disease and its Outcomes (TEMPO) 3:4 trial at doses of 60-120 mg/d. Blood samples were collected at baseline; at weeks 1, 2, and 3; at month 3; and every 3 months thereafter. RESULTS: In total, 134/135 (>99%) patients experienced ADRs. The most frequent ADRs were thirst (77.0%), pollakiuria (57.0%), polyuria (37.8%), and hyperuricemia (14.8%). Any unexpected ADRs were not reported in this trial. Most ADRs occurred early during treatment. Fourteen patients (10.4%) experienced hepatic events, and 8 (5.9%) experienced >3-fold increases above the upper limits of normal in serum alanine aminotransferase or aspartate aminotransferase levels between 3 and 9 months following tolvaptan initiation, which recovered after drug interruption. Of the 8 patients, 7 (5.2%) were previously allocated to the placebo arm in the TEMPO 3:4 trial and 4 (3.0%) discontinued due to the hepatic events. One patient (0.7%) was previously allocated to tolvaptan and experienced similar events in the TEMPO 3:4 trial. None of the hepatic ADRs met Hy's Law laboratory criteria. CONCLUSION: ADRs observed in this extension trial were similar to those identified in the TEMPO 3:4 trial and hepatic events were not progressive.

Ninety-day oral toxicity study of rice-derived γ-oryzanol in Sprague-Dawley rats.

A 90-day oral toxicity study of γ-oryzanol, a rice-derived triterpenoid ferulate, was performed by oral gavage administration to male and female Sprague-Dawley rats at doses of 0, 1000, and 2000 mg/kg body weight/day. All rats administered γ-oryzanol survived throughout the study period. Both male and female rats showed no toxicologically significant changes of the general signs, examination findings, body weight, food consumption, functional observational battery results, ophthalmological findings, urinalysis, hematology tests, clinical chemistry tests, organ weights, and necropsy findings. Moreover, there were no histopathological changes related to administration of γ-oryzanol in males and females from the 2000 mg/kg body weight/day group. In conclusion, the no observed adverse effect level (NOAEL) of γ-oryzanol exceeded 2000 mg/kg body weight/day for both male and female rats under the conditions of this study.

Identification and characterization of guanosine 5'-monophosphate reductase of Trypanosoma congolense as a drug target.

Trypanosoma congolense is one of the most prevalent pathogens which causes trypanosomosis in African animals, resulting in a significant economic loss. In its life cycle, T. congolense is incapable of synthesizing purine nucleotides via a de novo pathway, and thus relies on a salvage pathway to survive. In this study, we identified a gene from T. congolense, TcIL3000_5_1940, as a guanosine 5'-monophosphate reductase (GMPR), an enzyme that modulates the concentration of intracellular guanosine in the pathogen. The recombinant protein was expressed in Escherichia coli, and the gene product was enzymatically confirmed as a unique GMPR, designated as rTcGMPR. This enzyme was constitutively expressed in glycosomes at all of the parasite's developmental stages similar to other purine nucleotide metabolic enzymes. Mycophenolic acid (MPA) was found to inhibit rTcGMPR activity. Hence, it is a potential lead compound for the design of trypanocidal agents, specifically GMPR inhibitor.

Canagliflozin, a sodium glucose cotransporter 2 inhibitor, attenuates obesity-induced inflammation in the nodose ganglion, hypothalamus, and skeletal muscle of mice.

Chronic inflammation in systemic organs, such as adipose tissue, nodose ganglion, hypothalamus, and skeletal muscles, is closely associated with obesity and diabetes mellitus. Because sodium glucose cotransporter 2 (SGLT2) inhibitors exert both anti-diabetic and anti-obesity effects by promoting urinary excretion of glucose and subsequent caloric loss, we investigated the effect of canagliflozin, an SGLT2 inhibitor, on obesity-induced inflammation in neural tissues and skeletal muscles of mice. High-fat diet (HFD)-fed male C57BL/6J mice were treated with canagliflozin for 8 weeks. Canagliflozin attenuated the HFD-mediated increases in body weight, liver weight, and visceral and subcutaneous fat weight. Additionally, canagliflozin decreased blood glucose as well as the fat, triglyceride, and glycogen contents of the liver. Along with these metabolic corrections, canagliflozin attenuated the increases in the mRNA levels of the proinflammatory biomarkers Iba1 and Il6 and the number of macrophages/microglia in the nodose ganglion and hypothalamus. In the skeletal muscle of HFD-fed obese mice, canagliflozin decreased inflammatory cytokine levels, macrophage accumulation, and the mRNA level of the specific atrophic factor atrogin-1. Canagliflozin also increased the mRNA level of insulin-like growth factor 1, protected against muscle mass loss, and restored the contractile force of muscle. These findings suggested that SGLT2 inhibition disrupts the vicious cycle of obesity and inflammation, not only by promoting caloric loss, but also by suppression of obesity-related inflammation in both the nervous system and skeletal muscle.

Single-step enrichment of basophils from human peripheral blood by a novel method using a Percoll density gradient.

We have developed a novel continuous flow-through cell separation method using a Percoll density gradient. This method can continuously separate a large number of cells into five fractions according to their densities. To apply this method to the separation of basophils, Percoll density gradients were modified to improve basophil enrichment. When a set of Percoll density gradients was prepared (1.071, 1.075, 1.080, 1.084, and 1.090 g/mL) the basophils in a healthy volunteer were enriched by an average of 23.1 and 63.5% at Percoll densities of 1.075 (fraction 3) and 1.080 g/mL (fraction 4), respectively. On average, the yield of basophils was 1.66 × 10(5) cells in fraction 3 and 1.61 × 10(5) cells in fraction 4 from 9 mL of peripheral blood. The expression of CD203c (cluster of differentiation 203c) on separated basophils was upregulated by anti-immunoglobulin E stimulation similar to basophils in whole blood. Histamine release induced by calcium ionophore was also observed in the separated basophils. The present method will be useful for basophil enrichment since it preserves their function without using counterflow elutriation and immunological reagents, and this method will be effective as a preparative separation for cell purification by flow cytometry.

Mycophenolic Acid and Its Derivatives as Potential Chemotherapeutic Agents Targeting Inosine Monophosphate Dehydrogenase in Trypanosoma congolense.

This study aimed to evaluate the trypanocidal activity of mycophenolic acid (MPA) and its derivatives for Trypanosoma congolense The proliferation of T. congolense was completely inhibited by adding <1 µM MPA and its derivatives. In addition, the IMP dehydrogenase in T. congolense was molecularly characterized as the target of these compounds. The results suggest that MPA and its derivatives have the potential to be new candidates as novel trypanocidal drugs.