Anti-Inflammatory Effect of Korean Soybean Sauce (Ganjang) on Mice with Induced Colitis.

Inflammatory bowel disease (IBD), characterized by chronic inflammation of the gut, is caused by several factors. Among these factors, microbial factors are correlated with the gut microbiota, which produces short-chain fatty acids (SCFAs) via anaerobic fermentation. Fermented foods are known to regulate the gut microbiota composition. Ganjang (GJ), a traditional fermented Korean soy sauce consumed worldwide, has been shown to exhibit antioxidant, anticancer, anti-colitis, and antihypertensive activities. However, its effects on the gut microbiota remain unknown. In the present study, we aimed to compare the anti-inflammatory effects of GJ manufactured using different methods and investigate its effect on SCFA production in the gut. To evaluate the anti-inflammatory effects of GJ in the gut, we performed animal experiments using a mouse model of dextran sulfate sodium (DSS)-induced colitis. All GJ samples attenuated DSS-induced colitis symptoms, including reduced colonic length, by suppressing the expression of inflammatory cytokines. In addition, GJ administration modulated SCFA production in the DSS-induced colitis model. Overall, GJ exerted anti-inflammatory effects by reducing DSS-induced symptoms via regulation of inflammation and modulation of SCFA levels in a DSS-induced colitis model. Thus, GJ is a promising fermented food with the potential to prevent IBD.

Modified Thoracoabdominal Nerves Block Through Perichondral Approach: A Systematic Review and Meta-analysis.

PURPOSE: This systematic review and meta-analysis aimed to investigate the postoperative analgesic efficacy and safety of the modified thoracoabdominal nerve block through the perichondral approach (M-TAPA) in abdominal surgeries. DESIGN: Systematic review and meta-analysis. METHODS: We searched electronic databases to identify relevant studies comparing M-TAPA with conventional analgesic techniques. The primary outcome was the requirement for rescue analgesia at 12 and 24 hours postsurgery. Secondary outcomes included the 11-point numerical rating scale pain scores at 0, 1, 2, 4, 6, 8, 12, and 24 hours following surgery, global quality of recovery scores, and postoperative adverse events. FINDINGS: Five randomized controlled trials involving 308 patients were analyzed. M-TAPA showed no significant difference in the requirement for rescue analgesia at 12 hours (relative risk [RR]: 0.87; 95% confidence interval [CI]: 0.62, 1.22; P = .424; I2 = 40.7%; Ph = .185) and 24 hours (RR: 0.67; 95% CI: 0.22, 1.99; P = .252; I2 = 90.3%; Ph < .001) postsurgery compared to non-M-TAPA. No significant differences in numerical rating scale pain scores or global quality of recovery scores were found between the two groups (all P < .05). However, M-TAPA was associated with a lower occurrence of nausea (RR: 0.37; 95% CI: 0.22, 0.68; P < .001; I2 = 0%; Ph = .834), vomiting (RR: 0.32; 95% CI: 0.17, 0.62; P < .001; I2 = 0%; Ph = .884), and itching (RR: 0.38; 95% CI: 0.21, 0.70; P = .002; I2 = 0%; Ph = .826). CONCLUSIONS: There was no significant difference in analgesic efficacy and safety between M-TAPA and non-M-TAPA techniques.

Peripheral zone thickness in preoperative MRI is predictive of Trifecta achievement after Holmium laser enucleation of the prostate (HoLEP).

PURPOSE: To investigate various anatomical features of the prostate using preoperative MRI and patients' clinical factors to identify predictors of successful Holmium:YAG laser enucleation of the prostate (HoLEP). METHODS: 71 patients who had received HoLEP and undergone a 3.0-T prostate MRI scan within 6 months before surgery were retrospectively enrolled. MRI features (e.g., total prostate and transitional zone volume, peripheral zone thickness [PZT], BPH patterns, prostatic urethral angle, intravesical prostatic protrusion, etc.) and clinical data (e.g., age, body mass index, surgical technique, etc.) were analyzed using univariable and multivariable logistic regression to identify predictors of successful HoLEP. Successful HoLEP was defined as achieving the Trifecta, characterized by the contemporary absence of postoperative complications within 3 months, a 3-month postoperative maximum flow rate (Qmax) > 15 mL/s, and no urinary incontinence at 3 months postoperatively. RESULTS: Trifecta achievement at 3 months post-surgery was observed in 37 (52%) patients. Patients with Trifecta achievement exhibited a lower preoperative IPSS-quality of life score (QoL) (4.1 vs. 4.5, P = 0.016) and a thinner preoperative peripheral zone thickness (PZT) on MRI (7.9 vs.10.3 mm, P < 0.001). In the multivariable regression analysis, a preoperative IPSS-QoL score < 5 (OR 3.98; 95% CI, 1.21-13.07; P = 0.017) and PZT < 9 mm (OR 11.51; 95% CI, 3.51-37.74; P < 0.001) were significant predictors of Trifecta achievement after HoLEP. CONCLUSIONS: Alongside the preoperative QoL score, PZT measurement in prostate MRI can serve as an objective predictor of successful HoLEP. Our results underscore an additional utility of prostate MRI beyond its role in excluding concurrent prostate cancer.

A retrospective study for long-term oncologic and obstetric outcomes in cervical intraepithelial neoplasia treated with loop electrosurgical excision procedure: focus on surgical margin and human papillomavirus.

BACKGROUND: The present study aimed to evaluate the long-term oncological and obstetric outcomes following the loop electrosurgical excision procedure (LEEP) in patients with cervical intraepithelial neoplasia (CIN) and investigate the risk factors for recurrence and preterm birth. METHODS: This retrospective cohort study included patients who underwent LEEP for CIN 2-3 between 2011 and 2019. Demographic information, histopathological findings, postoperative cytology, and human papillomavirus (HPV) status were collected and analyzed. The Cox proportional hazards model and Kaplan-Meier curves with the log-rank test were used for risk factor analysis. RESULTS: A total of 385 patients treated with the LEEP were analyzed. Treatment failure, including recurrence or residual disease following surgery, was observed in 13.5% of the patients. Positive surgical margins and postoperative HPV detection were independent risk factors for CIN1 + recurrence or residual disease (HR 1.948 [95%CI 1.020-3.720], p = 0.043, and HR 6.848 [95%CI 3.652-12.840], p-value < 0.001, respectively). Thirty-one patients subsequently delivered after LEEP, and the duration between LEEP and delivery was significantly associated with preterm-related complications, such as a short cervix, preterm labor, and preterm premature rupture of the membrane (p = 0.009). However, only a history of preterm birth was associated with preterm delivery. CONCLUSIONS: Positive HPV status after LEEP and margin status were identified as independent risk factors for treatment failure in patients with CIN who underwent LEEP. However, combining these two factors did not improve the prediction accuracy for recurrence.

Effects of remimazolam combined with remifentanil on quality of recovery after ambulatory hysteroscopic surgery: a prospective, observational study.

BACKGROUND: Remimazolam, a new benzodiazepine, is known for its quick onset of effects and recovery time. Recently, it has been licensed for general anesthesia and sedation in Korea and its use is increasing in other countries. However, less is known about its effect on postoperative recovery. We used a patient-reported outcome questionnaire to examine the effect of remimazolam on postoperative recovery. METHODS: Patients who underwent hysteroscopy on day surgery basis were administered an induction dose of remimazolam 6 mg/kg/h followed by a maintenance dose of 1-2 mg/kg/h. After surgery, the translated Korean version of 15-item Quality of Recovery scale (QoR-15K) including post-discharge nausea and vomiting (PDNV) and/or pain, was surveyed 24 h after surgery to evaluate patient recovery. RESULTS: Total of 38 patients were enrolled in this prospective, observational study. All patients successfully completed QoR-15K. Only one patient scored low for moderate pain and PDNV. On average, patients scored 9 and above for all QoR-15K items except for moderate pain (8.66 ± 1.68). When QoR-15K items were grouped into dimensions, all dimensions scored an average of 9 or higher on a 10-point scale. In addition, 19 out of 38 patients gave score range of 148 to 150 out of possible 150. CONCLUSIONS: Psychometric evaluation based on postoperative QoR-15K among patients receiving remimazolam shows satisfactory patient recovery profiles without significant pain or PDNV. Considering its effectiveness and safety, remimazolam could be one of useful agents for general anesthesia of day surgery in terms of postoperative recovery.

Reduced side effects and improved pain management by continuous ketorolac infusion with patient-controlled fentanyl injection compared with single fentanyl administration in pelviscopic gynecologic surgery: a randomized, double-blind, controlled study.

BACKGROUND: A combination of opioids and adjunctive drugs can be used for intravenous patient-controlled analgesia (PCA) to minimize opioid-related side effects. We investigated whether two different analgesics administered separately via a dual-chamber PCA have fewer side effects with adequate analgesia than a single fentanyl PCA in gynecologic pelviscopic surgery. METHODS: This prospective, double-blind, randomized, and controlled study included 68 patients who underwent pelviscopic gynecological surgery. Patients were allocated to either the dual (ketorolac and fentanyl delivered by a dual-chamber PCA) or the single (fentanyl alone) group. Postoperative nausea and vomiting (PONV) and analgesic quality were compared between the two groups at 2, 6, 12, and 24 h postoperatively. RESULTS: The dual group showed a significantly lower incidence of PONV during postoperative 2-6 h (P = 0.011) and 6-12 h (P = 0.009). Finally, only two patients (5.7%) in the dual group and 18 (54.5%) in the single group experienced PONV during the entire postoperative 24 h and could not maintain intravenous PCA (odds ratio: 0.056, 95% CI [0.007, 0.229], P < 0.001). Despite the administration of less fentanyl via intravenous PCA during the postoperative 24 h in the dual group than in the single group (66.0 ± 77.8 vs. 383.6 ± 70.1 µg, P < 0.001), postoperative pain had no significant intergroup difference. CONCLUSIONS: Two different analgesics, continuous ketorolac and intermittent fentanyl bolus, administered via dual-chamber intravenous PCA, showed fewer side effects with adequate analgesia than conventional intravenous fentanyl PCA in gynecologic patients undergoing pelviscopic surgery.

Mesial temporal atrophy in preoperative MRI rather than steep Trendelenburg position is associated with postoperative delirium in patients undergoing a major urologic surgery.

PURPOSE: To investigate whether steep Trendelenburg in a major urologic surgery is associated with postoperative delirium, and to examine other potential clinical and radiologic factors predictive of postoperative delirium. METHODS: 182 patients who received a major urologic surgery and underwent a 3.0-T brain MRI scan within 1 year prior to the date of surgery were retrospectively enrolled. Preoperative brain MRIs were used to analyze features related to small vessel disease burden and mesial temporal atrophy. Presence of a significant mesial temporal atrophy was defined as Scheltens' scale ≥ 2. Patients' clinico-demographic data and MRI features were used to identify significant predictors of postoperative delirium using the logistic regression analysis. Independent predictors found significant in the univariate analysis were further evaluated in the multivariate analysis. RESULTS: Incidence of postoperative delirium was 6.0%. Patients with postoperative delirium had lower body mass index (21.3 vs. 25.0 kg/m2, P = 0.003), prolonged duration of anesthesia (362.7 vs. 224.7 min, P < 0.001) and surgery (302.2 vs. 174.5 min, P < 0.001), and had more significant mesial temporal atrophy (64% vs. 30%, P = 0.046). In the univariate analysis, female sex, type of surgery (radical prostatectomy over cystectomy), prolonged duration of anesthesia (≥ 6 h), and presence of a significant mesial temporal atrophy were significant predictors (all P-values < 0.050), but only the presence of significant mesial temporal atrophy was significant in the multivariate analysis [odds ratio (OR), 3.69; 95% CI 0.99-13.75; P = 0.046]. CONCLUSION: Steep Trendelenburg was not associated with postoperative delirium. Significant mesial temporal atrophy (Scheltens' scale ≥ 2) in preoperative brain MRI was predictive of postoperative delirium. TRIAL REGISTRATION: Not applicable.

Artificial intelligence model predicting postoperative pain using facial expressions: a pilot study.

PURPOSE: This study aimed to assess whether an artificial intelligence model based on facial expressions can accurately predict significant postoperative pain. METHODS: A total of 155 facial expressions from patients who underwent gastric cancer surgery were analyzed to extract facial action units (AUs), gaze, landmarks, and positions. These features were used to construct various machine learning (ML) models, designed to predict significant postoperative pain intensity (NRS ≥ 7) from less significant pain (NRS < 7). Significant AUs predictive of NRS ≥ 7 were determined and compared to AUs known to be associated with pain in awake patients. The area under the receiver operating characteristic curves (AUROCs) of the ML models was calculated and compared using DeLong's test. RESULTS: AU17 (chin raising) and AU20 (lip stretching) were found to be associated with NRS ≥ 7 (both P ≤ 0.004). AUs known to be associated with pain in awake patients did not show an association with pain in postoperative patients. An ML model based on AU17 and AU20 demonstrated an AUROC of 0.62 for NRS ≥ 7, which was inferior to a model based on all AUs (AUROC = 0.81, P = 0.006). Among facial features, head position and facial landmarks proved to be better predictors of NRS ≥ 7 (AUROC, 0.85-0.96) than AUs. A merged ML model that utilized gaze and eye landmarks, as well as head position and facial landmarks, exhibited the best performance (AUROC, 0.90) in predicting significant postoperative pain. CONCLUSION: ML models using facial expressions can accurately predict the presence of significant postoperative pain and have the potential to screen patients in need of rescue analgesia. TRIAL REGISTRATION NUMBER: This study was registered at ClinicalTrials.gov (NCT05477303; date: June 17, 2022).

Postoperative analgesic effects of the quadratus lumborum block in pediatric patients: a systematic review and meta-analysis.

Background: This study assessed the postoperative analgesic efficacy and safety of the quadratus lumborum block (QLB) in pediatric patients. Methods: Electronic databases were searched for studies comparing the QLB to conventional analgesic techniques in pediatric patients. The primary outcome was the need for rescue analgesia 12 and 24 hours after surgery. Secondary outcomes covered the Face-Legs-Activity-Cry-Consolability Scale (FLACC) scores at various time points; parental satisfaction; time to the first rescue analgesia; hospitalization time; block execution time; block failure rates, and adverse events. Results: Sixteen randomized controlled trials were analyzed involving 1,061 patients. The QLB significantly reduced the need for rescue analgesia both at 12 and 24 hours after surgery (12 hours, relative risk [RR]: 0.45; 95% confidence interval [CI]: 0.01, 0.88; 24 hours, RR: 0.51; 95% CI: 0.31, 0.70). In case of 24 hours after surgery, type 1 QLB significantly reduced the need for rescue analgesia (RR: 0.56; 95% CI: 0.36, 0.76). The QLB also exhibited lower FLACC scores at 1 hour (standardized mean difference [SMD]: -0.87; 95% CI: -1.56, -0.18) and 6 hours (SMD: -1.27; 95% CI: -2.33, -0.21) following surgery when compared to non-QLB. Among QLBs, type 2 QLB significantly extended the time until the first rescue analgesia (SMD: 1.25; 95% CI: 0.84, 1.67). No significant differences were observed in terms of parental satisfaction, hospitalization time, block execution time, block failure, or adverse events between QLB and non-QLB groups. Conclusions: The QLB provides non-inferior analgesic efficacy and safety to conventional methods in pediatric patients.

Effects of Cheonggukjang (Fermented Soybean) on the Development of Colitis-Associated Colorectal Cancer in Mice.

Colorectal cancer (CRC) is the third most common type of cancer and is caused by multiple factors. Chronic inflammation, known to cause inflammatory bowel disease (IBD), is closely associated with CRC. Cheonggukjang (CJ), a traditional Korean fermented soybean, is a functional food with anti-inflammatory effects in the intestines, but its anti-cancer effects have not yet been explored. In this study, we investigated the cancer-protective effects of cheonggukjang in an azoxymethane/DSS (AOM/DSS)-induced colitis-associated colorectal cancer (CAC) mouse model. The CJ alleviated AOM/DSS-induced pathological symptoms such as colonic shortening, increased spleen weight, tumor formation, and histological changes. It also modulated pro-inflammatory and anti-inflammatory cytokine levels via the suppression of NF-κB and inflammatory mediator signaling pathways. Furthermore, the CJ improved intestinal integrity by regulating mucin-associated and tight junction proteins. In addition, it suppressed tumor growth by regulating apoptosis and proliferation. These results highlight the anti-tumor effects of CJ in an AOM/DSS-induced CAC mouse model.

The Involvement of the Hypothalamus-Pituitary-Adrenal Axis in the Development of Hyperalgesia during the Early Postoperative Period.

INTRODUCTION: Hyperalgesia frequently occurs after surgery and is associated with adverse effects on surgical outcomes. Thus, we aimed to examine whether the hypothalamus-pituitary-adrenal (HPA) axis function after surgery is involved in the development of postoperative hyperalgesia. METHODS: Surgery- and pain-related variables were measured 24 and 48 h after the first and second total knee arthroplasties (TKAs) in postmenopausal patients undergoing 1-week-interval staged bilateral TKA. Two sets of saliva samples were consecutively collected from patients before (pre-T1) and 1 week after (post-T1) the first TKA (n = 69). HPA axis function was analyzed in a subgroup of 20 patients with a typical cortisol awakening response (CAR) in both the sets of saliva samples. RESULTS: Surgery-related variables were comparable between the first and second TKAs. However, pain-related variables (pain ratings and the amount of opioid analgesics consumed) were greater after the second than the first TKA. Cortisol and dehydroepiandrosterone (DHEA) secretion during the post-awakening period (CARauc and Daucawk, respectively) was higher at post-T1 than at pre-T1, but the molar CARauc/Daucawk ratio was comparable between the time points examined. No relationship was observed between the pre-T1 CARauc and pain ratings after the first TKA. However, post-T1 CARauc showed a positive correlation with pain ratings after the second TKA. Postoperative pain ratings were negatively correlated with Daucawk and positively correlated with the molar CARauc/Daucawk ratio at all examined time points. DISCUSSION/CONCLUSION: The results suggest that adrenocortical steroidogenic activity favoring the production of cortisol over DHEA after surgery may contribute to the development of hyperalgesia during the early postoperative period.

Controlling Gas Generation of Li-Ion Battery through Divinyl Sulfone Electrolyte Additive.

The focus of mainstream lithium-ion battery (LIB) research is on increasing the battery's capacity and performance; however, more effort should be invested in LIB safety for widespread use. One aspect of major concern for LIB cells is the gas generation phenomenon. Following conventional battery engineering practices with electrolyte additives, we examined the potential usage of electrolyte additives to address this specific issue and found a feasible candidate in divinyl sulfone (DVSF). We manufactured four identical battery cells and employed an electrolyte mixture with four different DVSF concentrations (0%, 0.5%, 1.0%, and 2.0%). By measuring the generated gas volume from each battery cell, we demonstrated the potential of DVSF additives as an effective approach for reducing the gas generation in LIB cells. We found that a DVSF concentration of only 1% was necessary to reduce the gas generation by approximately 50% while simultaneously experiencing a negligible impact on the cycle life. To better understand this effect on a molecular level, we examined possible electrochemical reactions through ab initio molecular dynamics (AIMD) based on the density functional theory (DFT). From the electrolyte mixture's exposure to either an electrochemically reductive or an oxidative environment, we determined the reaction pathways for the generation of CO2 gas and the mechanism by which DVSF additives effectively blocked the gas's generation. The key reaction was merging DVSF with cyclic carbonates, such as FEC. Therefore, we concluded that DVSF additives could offer a relatively simplistic and effective approach for controlling the gas generation in lithium-ion batteries.

Total intravenous anesthesia induced and maintained by a combination of remimazolam and remifentanil without a neuromuscular blocking agent: a prospective, observational pilot study.

BACKGROUND: A novel short-acting benzodiazepine, Remimazolam, has recently been approved for general anesthesia and sedation. Hence, we investigated the feasibility and safety of remimazolam during the induction and maintenance of general anesthesia without using a neuromuscular blocking agent (NMBA) in patients undergoing hysteroscopic surgery. METHODS: This prospective observational study included 38 patients undergoing hysteroscopic surgery. Remimazolam and remifentanil were the main anesthetic agents without an NMBA, and a supraglottic airway was inserted to protect the airway. The induction time, amount of each anesthetic agent used during anesthesia, intraoperative bispectral index (BIS) hemodynamic parameters, and recovery profiles were measured. RESULTS: General anesthesia was successfully administered to 37 patients using remimazolam and remifentanil without NMBA. The induction doses of remimazolam and remifentanil were 0.4 mg/kg (interquartile range [IQR] 0.34-0.47 mg/kg) and 1.07 µg/kg (IQR, 0.90-1.29 µg/kg), respectively. Additionally, the maintenance doses of remimazolam and remifentanil were 1.14 mg/kg/h (IQR, 0.88-1.55 mg/kg/h) and 0.06 µg/kg/min (IQR, 0.04-0.08 µg/kg/min), respectively. Intraoperative BIS values had risen temporarily > 60 in eight patients (21.6%) despite administration of 2 mg/kg/h of remimazolam; thus, they were treated with supplementary midazolam. The median recovery time was 7 min (IQR, 5-8 min) after 40 min (IQR, 40.0-57.5 min) of total mean anesthesia time. There was no correlation between the infusion dose of remimazolam and recovery profiles, such as recovery time, final BIS of anesthesia, modified observer assessment of alertness/sedation (OAA/S) scale or post-anesthesia recovery (PAR) score when arriving at the PACU, and length of stay in the PACU (all P > 0.05). CONCLUSION: Remimazolam can be combined with remifentanil without an NMBA in female patients who undergo hysteroscopic surgery, during which a supraglottic airway is a feasible method to protect the airway. TRIAL REGISTRATION: The study protocol was registered at ClinicalTrials.gov (NCT05025410) on 27/08/2021.

Effect of Detoxified Nano Sulfur Supplementation on the Growth, Nutrient Digestibility, Meat Quality, Excreta Microbes, Gas Emissions, and Blood Profiles of Broilers.

A 35-day experiment was conducted to evaluate the effects of the supplementation of mineral detoxified sulfur dispersion ((DSD); Patent No.: 10-1997773) on the growth performance, meat quality, excreta microbiota, gas emissions, nutrient digestibility, and blood profiles of broilers. In total, 720 one-day-old ROSS 308 broilers, with an initial body weight of 41.9±0.8 g, were divided into two (2) treatment groups with 20 replicate pens/groups composed of 18 birds per pen. Treatments consisted of 1) CON (the control), normal drinking water and 2) TRT (the treatment group), CON+0.001% DSD (1000:1 dilution ratio). Average daily feed intake (ADFI) and feed conversion ratio (FCR) increased in the TRT group (P<0.05) between days 1 to 7 and days 7 to 21 of the experimental period. Similarly, body weight gain (BWG) showed a significant increase (P<0.05) in the DSD-supplemented group throughout in the length of the experiment. With regard to meat quality, redness (a*) was higher, while drip loss was lower, on the 7th day in the DSD group. Furthermore, DSD supplementation increased (P<0.05) Lactobacillus excreta but decreased E. coli concentrations in the TRT group compared to the CON group. Notably, nutrient digestibility, excreta gas emission, and blood profiles did not show any significant differences (P>0.05). DSD supplementation, administered through drinking water, has a positive impact on the growth performance, meat quality, and excreta microbiota of broiler chickens.

Effect of Neuromuscular Blockade on Intraoperative Respiratory Mechanics and Surgical Space Conditions during Robot-Assisted Radical Prostatectomy: A Prospective Randomized Controlled Trial.

The aim of this study was to investigate whether deep neuromuscular blockade (NMB) may affect intraoperative respiratory mechanics, surgical condition, and recovery profiles in patients undergoing robot-assisted radical prostatectomy (RARP). Patients were randomly assigned to the moderate or deep NMB groups. Pneumoperitoneum was maintained with carbon dioxide (CO2) insufflation at 15 mmHg during surgery. The primary outcome was peak inspiratory pressure (PIP) after CO2 insufflation. Mean airway pressure (Pmean) and dynamic lung compliance (Cdyn) were also recorded. The surgeon rated the surgical condition and surgical difficulty on a five-point scale (1 = extremely poor; 2 = poor; 3 = acceptable; 4 = good; 5 = optimal). Recovery profiles, such as pulmonary complications, pain scores, and recovery time, were recorded. We included 58 patients in this study. No significant differences were observed regarding intraoperative respiratory mechanics including PIP, Pmean and Cdyn, between the two groups. The number of patients with optimal surgical conditions was significantly higher in the deep than in the moderate NMB group (29 vs. 20, p = 0.014). We found no differences in recovery profiles. In conclusion, deep NMB had no significant effect on the intraoperative respiratory mechanics but resulted in optimal endoscopic surgical conditions during RARP compared with moderate NMB.

Interplay of Enzyme Therapy and Dietary Management of Murine Homocystinuria.

Albeit effective, methionine/protein restriction in the management of classical homocystinuria (HCU) is suboptimal and hard to follow. To address unmet need, we developed an enzyme therapy (OT-58), which effectively corrected disease symptoms in various mouse models of HCU in the absence of methionine restriction. Here we evaluated short- and long-term efficacy of OT-58 on the background of current dietary management of HCU. Methionine restriction resulted in the lowering of total homocysteine (tHcy) by 38-63% directly proportional to a decreased methionine intake (50-12.5% of normal). Supplemental betaine resulted in additional lowering of tHcy. OT-58 successfully competed with betaine and normalized tHcy on the background of reduced methionine intake, while substantially lowering tHcy in mice on normal methionine intake. Betaine was less effective in lowering tHcy on the background of normal or increased methionine intake, while exacerbating hypermethioninemia. OT-58 markedly reduced both hyperhomocysteinemia and hypermethioninemia caused by the diets and betaine in HCU mice. Withdrawal of betaine did not affect improved metabolic balance, which was established and solely maintained by OT-58 during periods of fluctuating dietary methionine intake. Taken together, OT-58 may represent novel, highly effective enzyme therapy for HCU performing optimally in the presence or absence of dietary management of HCU.

Long-term uninterrupted enzyme replacement therapy prevents liver disease in murine model of severe homocystinuria.

Classical homocystinuria (HCU) is an inborn error of metabolism caused by loss of cystathionine ß-synthase (CBS) activity with the concomitant buildup of homocysteine. In knockout (KO) mice, a mouse model of HCU, complete lack of CBS is neonatally lethal. Administration of OT-58, an enzyme therapy for HCU, during the first 5 weeks of life rescued KO mice survival by preventing liver disease. Here, we studied the impact of a long-term uninterrupted OT-58 treatment or its absence beyond the neonatal period on liver pathology and metabolism. Plasma and liver metabolites of KO mice on OT-58 treatment were substantially improved or normalized compared with those receiving vehicle. Increased plasma activities of alanine aminotransferase and aspartate aminotransferase of vehicle-injected KO mice suggested the progression of liver damage with age and lack of treatment. At 3 months of age, liver histology showed no signs of hepatopathy in both vehicle- and OT-58-treated KO mice. However, moderate to severe liver disease, characterized by steatosis, hepatocellular necroses, disorganized endoplasmic reticulum, and swollen mitochondria, developed in 6-month-old vehicle-injected KO mice. KO mice on OT-58 treatment remained asymptomatic and were indistinguishable from age-matched healthy controls. Long-term uninterrupted OT-58 treatment was essential to prevent severe liver disease in the KO mouse model of HCU.

Behavior, body composition, and vascular phenotype of homocystinuric mice on methionine-restricted diet or enzyme replacement therapy.

Classic homocystinuria (HCU) is an inherited disorder characterized by elevated homocysteine (Hcy) in plasma and tissues resulting from cystathionine ß-synthase (CBS) deficiency. There is no cure, and patients are predominantly managed by methionine-restricted diet (MRD) to limit the production of Hcy. In this study, we used the I278T mouse model of HCU to evaluate the long-term impact of a novel enzyme replacement therapy [truncated human CBS C15S mutant modified with linear 20-kDa N-hydroxysuccinimide ester polyethylene glycol (OT-58)] on clinical end points relevant to human patients with HCU. In addition, we compared its efficacy on a background of either MRD or normal methionine intake [regular diet (REG)] to that of MRD alone. We found that, compared with untreated I278T mice, OT-58 treatment of I278T mice fed with the REG diet resulted in a 90% decrease in plasma Hcy concentrations and correction of learning/cognition, endothelial dysfunction, hemostasis, bone mineralization, and body composition. On background of the MRD, OT-58 performed equally well with plasma Hcy entirely normalized. The MRD alone decreased plasma Hcy by 67% and corrected the HCU phenotype in I278T mice. However, the MRD increased anxiety and reduced bone mineral content in both I278T mice and wild-type controls. This study shows that OT-58 is a highly efficacious novel treatment for HCU on the background of either normal or restricted methionine intake.-Majtan, T., Park, I., Cox, A., Branchford, B. R., di Paola, J., Bublil, E. M., Kraus, J. P. Behavior, body composition, and vascular phenotype of homocystinuric mice on methionine-restricted diet or enzyme replacement therapy.

Observation of acetyl phosphate formation in mammalian mitochondria using real-time in-organelle NMR metabolomics.

Recent studies point out the link between altered mitochondrial metabolism and cancer, and detailed understanding of mitochondrial metabolism requires real-time detection of its metabolites. Employing heteronuclear 2D NMR spectroscopy and 13C3-pyruvate, we propose in-organelle metabolomics that allows for the monitoring of mitochondrial metabolic changes in real time. The approach identified acetyl phosphate from human mitochondria, whose production has been largely neglected in eukaryotic metabolism since its first description about 70 years ago in bacteria. The kinetic profile of acetyl phosphate formation was biphasic, and its transient nature suggested its role as a metabolic intermediate. The method also allowed for the estimation of pyruvate dehydrogenase (PDH) enzyme activity through monitoring of the acetyl-CoA formation, independent of competing cytosolic metabolism. The results confirmed the positive regulation of mitochondrial PDH activity by p53, a well-known tumor suppressor. Our approach can easily be applied to other organelle-specific metabolic studies.

Pharmacokinetics and pharmacodynamics of PEGylated truncated human cystathionine beta-synthase for treatment of homocystinuria.

AIMS: PEGylated human truncated cystathionine beta-synthase, lacking the C-terminal regulatory domain (PEG-CBS), is a promising preclinical candidate for enzyme replacement therapy in homocystinuria (HCU). It was designed to function as a metabolic sink to decrease the severely elevated plasma and tissue homocysteine concentrations. In this communication, we evaluated pharmacokinetics (PK), pharmacodynamics (PD) and sub-chronic toxicity of PEG-CBS in homocystinuric mice, wild type rats and monkeys to estimate the minimum human efficacious dose for clinical trials. MAIN METHODS: Animal models received single or multiple doses of PEG-CBS. Activity of PEG-CBS and sulfur amino acid metabolites were determined in plasma and used to determine PK and PD. KEY FINDINGS: The plasma half-lives of PEG-CBS after a single subcutaneous (SC) injection were approximately 20, 44 and 73 h in mouse, rat and monkey, respectively. The SC administration of PEG-CBS resulted in a significant improvement or full correction of metabolic imbalance in both blood and tissues of homocystinuric mice. The PD of PEG-CBS in mouse was dose-dependent, but less than dose-proportional, with the maximal efficacy achieved at 8 mg/kg. PEG-CBS was well-tolerated in mice and monkeys, but resulted in dose-dependent minimal-to-moderate inflammation at the injection sites and vacuolated macrophages in rats. Allometric scaling of animal data was linear and the estimated human efficacious dose was determined as 0.66 mg/kg administered once a week. SIGNIFICANCE: These results provide critical preclinical data for the design of first-in-human PEG-CBS clinical trial.