The optimal time for laparoscopic excision of ovarian endometrioma: a prospective randomized controlled trial.

OBJECTIVE: This study aimed to explore the optimal time of laparoscopic cystectomy for unilateral ovarian endometrioma patients and evaluate the influence on ovarian reserve. MATERIALS AND METHODS: This prospective randomized controlled study included 88 women with unilateral ovarian endometrioma at a tertiary teaching hospital. All patients received their first identified diagnosis of ovarian endometrioma by ultrasound (> 4 cm and ≤ 10 cm) and were administered an oral contraceptive pill (OC) for one cycle before laparoscopy. They were randomly divided into two groups: laparoscopy at the late luteal phase (group LLP) (n = 44) (termination of OC for two days) and laparoscopy at the early follicular phase (group EFP) (n = 44) (day 3 after menstruation). Basic clinical characteristics were recorded. Serum Anti-Müllerian hormone (AMH) levels were measured at various times to predict ovarian reserve. Serum levels of Anti-Müllerian hormone (AMH) were measured at several time sites to predict the ovarian reserve; AMH and leukocyte esterase (LE) levels of the endometrioma wall were measured. RESULTS: Before surgery, serum AMH levels decreased in both groups from preoperative to one week and six months postoperatively. In contrast, the difference values of group EFP were larger than those of group LLP at postoperative one week and postoperative six months (1.87 ± 0.97 vs. 1.31 ± 0.93, P = 0.07; 1.91 ± 1.06 vs. 1.54 ± 0.93, P = 0.001). The mean rates of postoperative serum AMH decline were 37.92% and 46.34% in group EFP, significantly higher than those in group LLP (25.83% vs. 31.43%, P < 0.001). Ovarian endometrioma wall AMH of group LLP was significantly lower than that of group EFP ([22.86 ± 3.74] vs. [31.02 ± 5.23], P < 0.001). Meanwhile, ovarian endometrioma LE concentration of group LLP was significantly higher than that of group EFP ([482.83 ± 115.88] vs. [371.68 ± 84.49], P<0.001). There was also a significant inverse correlation between leukocyte esterase and AMH concentration in an ovarian endometrioma cyst wall (r=-0.564, P<0.001). CONCLUSION(S): The optimal time for laparoscopic cystectomy for patients with first identified unilateral ovarian endometrioma is the late luteal phase, which reduces ovarian tissue loss and preserves ovarian reserve effectively and safely.

The Application Value of Contrast-Enhanced Ultrasonography in Assessing the Efficacy of Ultrasound-Guided Lauromacrogol Injections in Cesarean Scar Pregnancies.

OBJECTIVE: To investigate the application value of contrast-enhanced ultrasonography (CEUS) in ultrasound-guided lauromacrogol injections in patients with cesarean scar pregnancies (CSP). METHODS: A total of 31 patients diagnosed with CSP, who underwent an ultrasound-guided lauromacrogol injection + curettage in our hospital between February 2019 and December 2020 and had a complete recovery confirmed by a postoperative ultrasound review and serum ß-human chorionic gonadotropin (ß-hCG) assay, were enrolled as the study subjects. According to the volume of intraoperative blood loss and the duration of postoperative vaginal bleeding, the patients were divided into two groups, with 19 in the significantly effective group (Group A) and 12 in the effective group (Group B). The recorded clinical data, including age, duration of amenorrhea, number of pregnancies, number of deliveries, time since last cesarean delivery, number of cesarean deliveries, and preoperative serum ß-hCG levels, were retrospectively analyzed. The morphological indicators in CEUS before the lauromacrogol injection, as well as immediately and 12-24 hours after the injection, were compared between the groups. RESULTS: In Group A, the post-injection CEUS showed no enhancement, single strip enhancement, and sparse punctate enhancement, while in Group B, it showed a more irregular ring and local patch enhancement. In addition, the number of cases where the CEUS showed no enhancement 12-24 hours after the injection was more than that of the immediate CEUS after the injection. In Group A, four (21.1%) cases showed a single strip-like blood flow on the immediate postoperative CEUS, four (21.1%) cases showed a sparsely dotted blood flow on the immediate postoperative CEUS, and three cases turned into no enhancement 12-24 hours after the injection. A total of four cases in Group B showed that the contrast enhancement range 12-24 hours after the injection was reduced compared with that of the immediate contrast after the injection. CONCLUSION: Contrast-enhanced ultrasonography can guide the location selection of the lauromacrogol injection in patients with CSP, and its postoperative morphological indicators can adequately predict the therapeutic effect after curettage and guide clinical management.

Preliminary hysteroscopic tubal hydrotubation improves fertility outcomes after laparoscopic salpingotomy for tubal ampullary pregnancy.

BACKGROUND: Salpingotomy may change the anatomical structure and patency of the fallopian tube, which may affect the fallopian function. This study is to investigate the clinical efficacy of preliminary hysteroscopic tubal hydrotubation (HTH) after laparoscopic salpingotomy for tubal ampullary pregnancy. METHODS: A retrospective study was carried out, 140 women underwent laparoscopic salpingotomy for ampullary pregnancy from March 2013 to March 2017. Some patients received HTH in the 1st month and hysterosalpingography (HSG) in the 3rd month after salpingotomy (HTH group; n = 95), and some patients only received HSG in the 3rd month after salpingotomy (control group; n = 45). Clinical data, tubal patency and fertility outcome were evaluated after follow-up of 15 months. RESULTS: The tubal patency rate of the operated side was significantly higher in the HTH group than that in the control group (89.47% vs 68.89%, P < 0.05). The intrauterine pregnancy (IUP) rate was significantly higher in the HTH group (76.47% vs 51.11%, P < 0.05), and the recurrent ectopic pregnancy rate in the operated side was significantly lower in the HTH group than in the control group (9.41% vs 22.22%, P < 0.05). Logistic regression analysis showed that the positive factor for IUP was HTH (OR = 3.109, 95% CI 1.439-6.714, P = 0.004), while the negative factors were history of pelvic inflammatory disease (PID) (OR = 0.167, 95% CI 0.074-0.377, P < 0.001) and history of tubal infertility (OR = 0.286, 95% CI 0.113-0.723, P < 0.05). CONCLUSION: Preliminary HTH after laparoscopic salpingotomy for ampullary pregnancy could improve reproductive function and lead to a better fertility outcome. Patients without history of PID or tubal infertility may be the most suitable ones for HTH after salpingotomy.

Crops For the Future (CFF): an overview of research efforts in the adoption of underutilised species.

MAIN CONCLUSION: Crops For the Future (CFF), as an entity, has established a broad range of research activities to promote the improvement and adoption of currently underutilised crops. This paper summarises selected research activities at Crops For the Future (CFF) in pursuit of its mission 'to develop solutions for diversifying future agriculture using underutilised crops'. CFF is a research company focussed on the improvement of underutilised crops, so that they might be grown and consumed more widely with benefits to human food and nutritional security; its founding guarantors were the Government of Malaysia and the University of Nottingham. From its base in Malaysia, it engages in research around the world with a focus on species and system diversification. CFF has adopted a food system approach that adds value by delivering prototype food, feed and knowledge products. Bambara groundnut (Vigna subterranea) was adopted as an exemplar crop around which to develop CFF's food system approach with emphasis on the short-day photoperiod requirement for pod-filling and the hard-to-cook trait. Selective breeding has allowed the development of lines that are less susceptible to photoperiod but also provided a range of tools and approaches that are now being exploited in other crops such as winged bean (Psophocarpus tetragonolobus), amaranth (Amaranthus spp.), moringa (Moringa oleifera) and proso (Panicum miliaceum) and foxtail (Setaria italica) millets. CFF has developed and tested new food products and demonstrated that several crops can be used as feed for black soldier fly which can, in turn, be used to feed fish thereby reducing the need for fishmeal. Information about underutilised crops is widely dispersed; so, a major effort has been made to develop a knowledge base that can be interrogated and used to answer practical questions about potential exploitation of plant and nutritional characteristics. Future research will build on the success with Bambara groundnut and include topics such as urban agriculture, rural development and diversification, and the development of novel foods.

Experiences of Singapore nurses as second victims: A qualitative study.

Nurses, like other healthcare professionals, such as physicians, pharmacists, and therapists, are susceptible to unanticipated patient harm, in which they suffer as second victims due to the immense personal and professional impact from the adverse event. The aim of the present study was to explore the psychological responses, coping strategies, and support needs of Singapore nurses as second victims of adverse events. A descriptive qualitative study was adopted. Eight participants (6 women and 2 men) who had been involved in an adverse event were interviewed and audio-recorded. Thematic analysis was performed to analyze the data. Seven themes emerged from the thematic analysis: responding psychologically after the event, feeling others' prejudice, having intrusive thoughts, drawing valuable lessons from the event, coping to recover after the event, taking responsibility for the mistakes made, and finding self-identity. The harmful effects of adverse events on nurses are long lasting. Second-victim nurses adopted various coping strategies to recover. The findings from the present study will guide the development of effective second-victim support programs.

Association between blood cadmium levels and malnutrition in peritoneal dialysis.

BACKGROUND: Malnutrition is associated with an increased risk of cardiovascular death and may cause protein-energy wasting in individuals with chronic kidney disease. A previous study demonstrated that blood cadmium levels (BCLs) were associated with malnutrition in maintenance hemodialysis (MHD) patients. However, the correlation between cadmium exposure and malnutrition remains unclear in chronic peritoneal dialysis (CPD) patients. This study examined the possible adverse effects of environmental cadmium exposure in CPD patients. METHODS: A total of 301 CPD patients were enrolled and divided into 3 study groups based on the following BCL tertiles: low (<0.19 µg/L), middle (0.19-0.39 µg/L), and high (>0.39 µg/L). Demographic, hematological, biochemical, and dialysis-related data were obtained for analysis. The analysis also included values of nutritional and inflammatory markers. RESULTS: The BCLs of CPD patients were lower than those of MHD patients. At baseline, patients in the high BCL group were older and had a higher prevalence of diabetes mellitus but lower serum albumin, creatinine, and phosphate levels than the patients in the other 2 groups. After adjusting for potential variables, stepwise backward multiple linear regression analysis revealed that age and alanine aminotransferase levels were positively associated with logarithmic transformation of BCLs (log BCLs), while serum albumin levels were negatively associated with log BCLs in CPD patients. The log BCLs were a significant determinant (beta coefficient ± standard error = -0.185 ± 0.074; P = 0.013) of nutritional status and significantly associated with the presence of malnutrition (odds ratio = 2.64; 95% confidence interval: 1.07-6.48; P = 0.035) in CPD patients after adjustment for related variables. CONCLUSIONS: BCL is significantly associated with nutritional status and malnutrition in CPD patients. Therefore, it is important for CPD patients to avoid environmental exposure to cadmium such as through smoking and consumption of cadmium-rich foods.

Biochemical characterization and mutational analysis of the NurA protein from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5.

Archaeal NurA protein plays a key role in producing 3'-single stranded DNA used for homologous recombination repair, together with HerA, Mre11, and Rad50. Herein, we describe biochemical characteristics and roles of key amino acid residues of the NurA protein from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5 (Tba-NurA). Tba-NurA possesses 5'-3' exonuclease activity for degrading DNA, displaying maximum efficiency at 45 °C-65 °C and at pH 8.0 in the presence of Mn2+. The thermostable Tba-NurA also possesses endonuclease activity capable of nicking plasmid DNA and circular ssDNA. Mutational data demonstrate that residue D49 of Tba-NurA is essential for exonuclease activity and is involved in binding ssDNA since the D49A mutant lacked exonuclease activity and reduced ssDNA binding. The R96A and R129A mutants had no detectable dsDNA binding, suggesting that residues R96 and R129 are important for binding dsDNA. The abolished degradation activity and reduced dsDNA binding of the D120A mutant suggest that residue D120 is essential for degradation activity and dsDNA binding. Additionally, residues Y392 and H400 are important for exonuclease activity since these mutations resulted in exonuclease activity loss. To our knowledge, it is the first report on biochemical characterization and mutational analysis of the NurA protein from Thermococcus.

Education level is associated with mortality in male patients undergoing maintenance hemodialysis.

BACKGROUND: Studies of the correlation between education levels and mortality in hemodialysis (HD) patients are rare. The aim of this multi-center study was to investigate the relationship between education levels and 3-year mortality rates in HD patients. METHODS: A total of 935 HD patients from 3 HD centers participated in this 3-year prospective observational study. Education levels were categorized as either less than senior high school and above or equal to senior high school. The causes of death and mortality rates were also analyzed for each subgroup. RESULTS: At the end of the 3-year follow-up period, 164 patients had died. In the male group, forward stepwise Cox regression analysis revealed that age, HD duration, hypertension, creatinine level, serum albumin level ≥3.6 g/dl, anuria, Kt/Vurea, and high education level were significant predictive factors for 3-year mortality rates. CONCLUSION: This prospective observational study demonstrated that education level was associated with mortality in men undergoing HD.

Death-related factors of systemic lupus erythematosus patients associated with the course of disease in Chinese populations: multicenter and retrospective study of 1,958 inpatients.

Despite the improved survival rate among systemic lupus erythematosus (SLE) patients, there are many factors associated with the mortality of SLE. In the current study, death-related factors of patients associated with course of disease were surveyed. Retrospective study was used. Mortalities among these three groups (group A, B and C, the course of disease was ≤ 5 years, 5-10 years and > 10 years, respectively) were calculated and compared. Various factors related to mortality were analyzed. Male SLE patients died relatively more than female patients. The total mortality was 8.5 %. The mortalities were significant difference in group A, B and C which were 9.4, 4.8 and 8.9 %, respectively. The mortalities of group A and group C were significantly higher than that of group B, but there was no significant difference between mortalities of group A and group C. The most common death-related factor was infection, followed by involved disorders in renal, brain, multisystem, heart, etc. The mortalities resulted from neuropsychiatric systemic lupus erythematosus (NPSLE), pulmonary infection, involved digestive system and hematological system were significantly different between three groups. There was no difference between mortalities of group A and group C associated with pulmonary infection and NPSLE. Patients in group C died more than in group A from involved renal, heart, multisystem, etc, while group A had more patients than group C who died of pulmonary infection, involved hematological system. In conclusion, Male SLE patients have worse outcome than female patients. Infection and active SLE are not only contributors to the death of early stage patients, but also to that of later stage patients.

Biochemical characterization and mechanistic insight of the family IV uracil DNA glycosylase from Sulfolobus islandicus REY15A.

Uracil DNA glycosylase (UDG) can remove uracil from DNA, thus playing an essential role in maintaining genomic stability. Family IV UDG members are mostly widespread in hyperthermophilic Archaea and bacteria. In this work, we characterized the family IV UDG from the hyperthermophilic crenarchaeon Sulfolobus islandicus REY15A (Sis-UDGIV) biochemically, and dissected the roles of nine conserved residues in uracil excision by mutational analyses. Biochemical data demonstrate that Sis-UDGIV displays maximum efficiency for uracil excision at 50 °C ~ 70 °C and at pH 7.0-9.0. Additionally, the enzyme has displays a weak activity without a divalent metal ion, but maximum activity with Mg2+. Our mutational analyses show that residues E48 and F55 in Sis-UDGIV are essential for uracil removal, and residues E48, F55, R87, R92 and K146 are responsible for binding DNA. Importantly, we systemically revealed the roles of four conserved cysteine residues C14, C17, C86 and C102 in Sis-UDGIV that are required for being ligands of FeS cluster in maintaining the overall protein conformation and stability by circular dichroism analyses. Overall, our work has provided insights into biochemical function and DNA-binding specificity of archaeal family IV UDGs.

Shifts and importance of viable bacteria in treatment of DSS-induced ulcerative colitis mice with FMT.

Background and Aims: Ulcerative colitis (UC) has become a global public health concern, and is in urgent need of novel therapies. Fecal microbiota transplantation (FMT) targeting gut microbiota has recently been applied to the treatment of UC. Despite its recent successes, it is still largely unknown how FMT functionally modulates the gut microbiota and improves the disease. Methods: We prospectively collected fecal samples from the 40 mice (30 mice for dextran sulfate sodium (DSS)-induced, 10 for controls), followed by Propidium monoazide treatment for 16S rRNA gene sequencing. These 30 mice were divided equally into 3 groups, which were transplanted with original donor microbiota (DO), inactivated donor microbiota (DI) and saline, respectively. Subsequently, we used 16S rRNA gene sequencing to analyze the viable gut bacteria of ulcerative colitis (UC) mice and histological analysis to evaluate the effects of fecal microbiota transplantation (FMT) with viable microbiota. Results: We demonstrated that the community structure of viable bacteria was significantly different from fecal bacteria based on total DNA. Furthermore, the intestinal viable microbiota and colonic mucosal structure of mice were significantly changed by DSS induction. The histological analysis showed that only the mice treated with original donor microbiota group (HF) achieved a significant improvement. Compared with inactivated donor microbiota group (IF) and saline (NF), Lactobacillus and Halomonas were significantly enriched in the HF group. Conclusion: We inferred that only live bacteria from human donor reversed the histopathology and symptoms of UC in mice and altered the gut microbiota. The activity of gut microbiota in donor samples should be considered in FMT and that detailed analysis of viable microbiota is essential to understand the mechanisms by which FMT produces therapeutic effects in the future.

Effect of chelation therapy on progressive diabetic nephropathy in patients with type 2 diabetes and high-normal body lead burdens.

BACKGROUND: A previous study in type 2 diabetic patients with high-normal body lead burdens showed that EDTA chelation therapy for 3 months slows progressive diabetic nephropathy during a 12-month follow-up. The effect of a longer course of therapy on kidney function decrease over a longer follow-up is not known. STUDY DESIGN: A 12-month run-in phase, then a randomized single-blind study with a 27-month intervention. SETTING & PARTICIPANTS: University medical center; 50 patients (serum creatinine, 1.5-3.9 mg/dL) with high-normal body lead burden (≥80-<600 µg) were randomly assigned to the treatment and control groups. INTERVENTION: The treatment group received weekly chelation therapy for 3 months to reduce their body lead burden to <60 µg and then as needed for 24 months to maintain this level. The control group received placebo for 3 months and then weekly for 5 weeks at 6-month intervals for 24 months. OUTCOMES: The primary end point was change in estimated glomerular filtration rate (eGFR) over time. A secondary end point was a 2-fold increase in baseline serum creatinine level or the requirement for renal replacement therapy. MEASUREMENTS: Body lead burdens were assessed by EDTA mobilization tests and eGFR was calculated using the equation for Chinese patients with type 2 diabetes. RESULTS: Mean baseline eGFRs in the treatment and control groups were similar. After 3 months of chelation therapy, the change in eGFR in the treatment group (+1.0 ± 4.8 mL/min/1.73 m(2)) differed significantly from that in the control group (-1.5 ± 4.8 mL/min/1.73 m(2); P = 0.04). In the subsequent 24-month intervention, the yearly rate of decrease in eGFR (5.6 ± 5.0 mL/min/1.73 m(2) per year) in the treatment group was slower than that (9.2 ± 3.6 mL/min/1.73 m(2) per year; P = 0.04) in the control group. 17 (68%) control-group patients and 9 (36%) treatment-group patients achieved the secondary end point. LIMITATIONS: Small sample size, not double blind. CONCLUSIONS: A 27-month course of EDTA chelation therapy retards the progression of diabetic nephropathy in type 2 diabetic patients with high-normal body lead burdens.

Lead level in seminal plasma may affect semen quality for men without occupational exposure to lead.

BACKGROUND: Infertility affects approximately 10-15% of reproductive-age couples. Poor semen quality contributes to about 25% of infertile cases. Resulting from the direct effect on testicular function or hormonal alterations, heavy metals exposure has been related to impaired semen quality. The objective of this study was to assess the level of lead in the seminal plasma in men without occupational exposure to lead, and to determine the relationship between semen quality and lead concentration in the semen. METHODS: This is a prospective and nonrandomized clinical study conducted in University infertility clinic and academic research laboratory. Three hundred and forty-one male partners of infertile couples undergoing infertility evaluation and management were recruited to the study. Semen samples collected for the analyses of semen quality were also used for the measurement of lead concentrations. Semen samples were evaluated according to the WHO standards. RESULTS: All subjects were married and from infertile couples without occupational exposure to lead. There is a significant inverse correlation between the lead concentration in seminal plasma and sperm count. A higher semen lead concentration was correlated with lower sperm count, but not with semen volume, sperm motility or sperm morphology as assessed by simple linear regression. CONCLUSIONS: We found that semen lead concentration was significantly higher among the patients with lower sperm count. To our knowledge, this is the first study to demonstrate that a high level of lead accumulation in semen may reduce the sperm count contributing to infertility of men without occupational exposure to lead.

Spectrum of toxic hepatitis following intentional paraquat ingestion: analysis of 187 cases.

INTRODUCTION: This retrospective observational study examined the clinical features, the degrees of toxic hepatitis, physiological markers and clinical outcomes after intentional paraquat poisoning and sought to determine what association, if any, might exist between these findings. METHODS: A total of 187 patients were referred for management of intentional paraquat ingestion between 2000 and 2010. Patients were categorized into two groups according to their hepatic complication, i.e. with (N = 87) or without (N = 100) toxic hepatitis. Demographic, clinical and laboratory data were obtained for analysis. Mortality rates were also analysed. RESULTS: It was found that patients with toxic hepatitis were younger (39.7 ± 13.7 vs 44.2 ± 16.6 year old, P = 0.046), and suffered from greater incidences of acute respiratory failure (63.2 vs 48.0%, P = 0.037) and acute renal failure (75.9 vs 56.0%, P = 0.004) than patients without hepatitis. The hospitalization period was longer in patients with hepatitis than without hepatitis (16.2 ± 14.6 vs 11.2 ± 12.1 days, P = 0.012), even though there was no difference in mortality rate between both groups (56.3 vs 53.0%, P = 0.649). Notably, the symptoms of toxic hepatitis developed within 6.7 ± 6.3 days of exposure to paraquat with aspartate aminotransferase (AST) 138 ± 156 U/L, alanine aminotransferase (ALT) 127 ± 114 U/L and total bilirubin 2.7 ± 2.6 mg/dL. The hepatitis peaked at 9.5 ± 8.8 days with AST 125 ± 139 U/L, ALT 183 ± 181 U/L and total bilirubin 3.2 ± 3.6 mg/dL. Nevertheless, the symptoms resolved within 17.3 ± 9.8 days of paraquat exposure, and none of the patients died of hepatic complication. CONCLUSIONS: A substantial proportion of paraquat patients suffered from hepatic complication (46.52%), but the spectrum of hepatitis in these patients seemed mild and transient.

Biochemical and functional characterization of a thermostable RecJ exonuclease from Thermococcus gammatolerans.

RecJ is ubiquitous in bacteria and Archaea, and play an important role in DNA replication and repair. Currently, our understanding on biochemical function of archaeal RecJ is incomplete due to the limited reports. The genome of the hyperthermophilic and radioresistant euryarchaeon Thermococcus gammatolerans encodes one putative RecJ protein (Tga-RecJ). Herein, we report biochemical characteristics and catalytic mechanism of Tga-RecJ. Tga-RecJ can degrade ssDNA in the 5'-3' direction at high temperature as observed in Thermococcus kodakarensis RecJ and Pyrococcus furiosus RecJ, the two closest homologs of the enzyme. In contrasted to P. furiosus RecJ, Tga-RecJ lacks 3'-5' ssRNA exonuclease activity. Furthermore, maximum activity of Tga-RecJ is observed at 50 °C ~ 70 °C and pH 7.0-9.0 with Mn2+, and the enzyme is the most thermostable among the reported RecJ proteins. Additionally, the rates for hydrolyzing ssDNA by Tga-RecJ were estimated by kinetic analyses at 50 °C ~ 70 °C, thus revealing its activation energy (10.5 ± 0.6 kcal/mol), which is the first report on energy barrier for ssDNA degradation by RecJ. Mutational studies showed that the mutations of residues D36, D83, D105, H106, H107 and D166 in Tga-RecJ to alanine almost completely abolish its activity, thereby suggesting that these residues are essential for catalysis.

Biochemical characterization and mutational analysis of a mismatch glycosylase from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5.

Mismatch glycosylase (MIG) can excise thymine and uracil from mutagenic T:G and U:G mispairs, which arise from cytosine and 5-methylcytosine deamination, respectively. Here, we present evidence that a thermostable MIG from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5 (Tb-MIG) can remove thymine and uracil from T:G and U:G mispairs at high temperature, albeit at a low efficiency for U:G mispair. The enzyme displays maximum efficiency at 70 oC - 75 °C and pH 7.0-8.0. Tb-MIG is extremely thermostable, retaining 50% activity after heating at 100 oC for 2 hrs. In addition, Tb-MIG is a bifunctional glycosylase with an AP lyase activity, then resembles the MIG from the hyperthermopilic crenarchaeon Pyrobaculum aerophilium, but contrasts with the MIG from the hyperthermopilic crenarchaeon Aeropyrum pernix. Importantly, we show that residues Y133 and D151 in Tb-MIG are essential for thymine removal, and that residues A58, N153 and R156 are involved in thymine removal. Compared with the wild-type protein, the A58D and Y133K mutants display the increased AP lyase activity, confirming the essential roles played by the correspondingly conserved Asp and Lys in endonuclease III for AP site cleavage. Overall, our work is the first biochemical characterization of a hypthermophilic euryarchaeal MIG, augmenting our understanding on archaeal MIG function.

Biochemical characterization and mutational analysis of a novel flap endonuclease 1 from Thermococcus barophilus Ch5.

Flap endonuclease 1 (FEN1) plays important roles in DNA replication, repair and recombination. Herein, we report biochemical characteristics and catalytic mechanism of a novel FEN1 from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5 (Tb-FEN1). As expected, the recombinant Tb-FEN1 can cleave 5'-flap DNA. However, the enzyme has no activity on cleaving pseudo Y DNA, which sharply contrasts with other archaeal and eukaryotic FEN1 homologs. Tb-FEN1 retains 24% relative activity after heating at 100 °C for 20 min, demonstrating that it is the most thermostable among all reported FEN1 proteins. The enzyme displays maximal activity in a wide range of pH from 7.0 to 9.5. The Tb-FEN1 activity is dependent on a divalent metal ion, among which Mg2+ and Mn2+ are optimal. Enzyme activity is inhibited by NaCl. Kinetic analyzes estimated that an activation energy for removal of 5'-flap from DNA by Tb-FEN1 was 35.7 ± 4.3 kcal/mol, which is the first report on energy barrier for excising 5'-flap from DNA by a FEN1 enzyme. Mutational studies demonstrate that the K87A, R94A and E154A amino acid substitutions abolish cleavage activity and reduce 5'-flap DNA binding efficiencies, suggesting that residues K87, R94, and E154 in Tb-FEN1 are essential for catalysis and DNA binding as well. Overall, Tb-FEN1 is an extremely thermostable endonuclease with unusual features.

An Infant Formula with Large, Milk Phospholipid-Coated Lipid Droplets Supports Adequate Growth and Is Well-Tolerated in Healthy, Term Asian Infants: A Randomized, Controlled Double-Blind Clinical Trial.

Lipids are essential for healthy infant growth and development. The structural complexity of lipids in human milk is not present in infant milk formula (IF). A concept IF was developed mimicking more closely the structure and composition of human milk fat globules. The current study evaluates whether a concept IF with large, milk phospholipid-coated lipid droplets (mode diameter 3 to 5 µm) is equivalent to standard IF with regard to growth adequacy and safety in healthy, term Asian infants. In this randomized, double-blind, controlled trial, infants were randomized after parents decided to introduce formula. Infants received a standard IF with (Control) or without the specific prebiotic mixture scGOS/lcFOS (9:1 ratio; Control w/o prebiotics), or a Concept IF with large, milk phospholipid-coated lipid droplets and the prebiotic mixture. A group of 67 breastfed infants served as a reference. As a priori defined, only those infants who were fully intervention formula-fed ≤28 days of age were included in the equivalence analysis (Control n = 29; Control w/o prebiotics n = 28; Concept n = 35, per-protocol population). Primary outcome was daily weight gain during the first four months of life, with the difference between the Concept and Control as the key comparison of interest. Additionally, adverse events, growth and tolerance parameters were evaluated. Equivalence of daily weight gain was demonstrated between the Concept and Control group after additional correction for ethnicity and birthweight (difference in estimated means of 0.1 g/d, 90%CI [-2.30, 2.47]; equivalence margin +/- 3 g/d). No clinically relevant group differences were observed in secondary growth outcomes, tolerance outcomes or number, severity or relatedness of adverse events. This study corroborates that an infant formula with large, milk phospholipid-coated lipid droplets supports adequate growth and is well tolerated and safe for use in healthy infants.

Evaluation of the clinical utility of the PromarkerD in-vitro test in predicting diabetic kidney disease and rapid renal decline through a conjoint analysis.

BACKGROUND: Early identification of patients at risk of developing diabetic kidney disease or rapid renal decline is imperative for appropriate patient management, but traditional methods of predicting renal decline are limited. OBJECTIVE: This study evaluated the impact of PromarkerD, a biomarker-based blood test predicting the risk of diabetic kidney disease (DKD) and rapid renal decline. METHODS: Conjoint analysis clarified the importance of PromarkerD and other patient attributes to physician decisions for type 2 diabetes patients. Forty-two patient profiles were generated, with varying levels of albuminuria, estimated glomerular filtration rate (eGFR), blood pressure, hemoglobin A1c (HbA1c), age, and PromarkerD result. A web-based survey asked each physician to make monitoring/treatment decisions about eight randomly selected profiles. Data were analyzed using multivariable logit models. RESULTS: Two hundred three primary care physicians and 197 endocrinologists completed the survey. PromarkerD result was most important for increasing the frequency of risk factor monitoring. PromarkerD was second to HbA1c in importance for deciding to prescribe sodium/glucose cotransporter-2 inhibitors (SGLT2s) with a DKD indication, second to blood pressure for increasing the dose of lisinopril, and second to eGFR for replacing ibuprofen with a non-nephrotoxic medication. Compared with no PromarkerD results, a high-risk PromarkerD result was associated with significantly higher odds of increasing monitoring frequency (odds ratio [OR]: 2.56, 95% confidence interval: 1.90-3.45), prescribing SGLT2s (OR: 1.98 [1.56-2.52]), increasing lisinopril dose (OR: 1.48 [1.17-1.87]), and replacing ibuprofen (OR: 1.78 [1.32-2.40]). A low-risk PromarkerD result was associated with significantly lower odds of increasing monitoring frequency (OR: 0.48 [0.37-0.64]), prescribing SGLT2s (OR: 0.70 [0.56-0.88]), and replacing ibuprofen (OR: 0.75 [0.57-0.99]). CONCLUSION: PromarkerD could increase adoption of renoprotective interventions in patients at high risk for renal decline and lower the likelihood of aggressive treatment in those at low risk. Further studies are needed to assess patient outcomes with PromarkerD in real-world practice.