Relationship between faecal metronidazole and lactoferrin concentrations to clinical response of patients with Clostridioides difficile.

OBJECTIVES: We investigated patients with Clostridioides difficile-associated diarrhoea to see if clinical resolution correlated with faecal concentrations of metronidazole or markers of inflammation. METHODS: Faecal metronidazole, lactoferrin and serum CRP were measured daily. These were then compared with clinical progress. RESULTS: Metronidazole concentration correlated with lactoferrin (ρ = 0.17, p = 0.015), CRP (ρ = 0.23, p < 0.001) and number of diarrhoeal stools per day (ρ = 0.29, p < 0.001). Lactoferrin correlated with CRP (ρ = 0.57, p < 0.001) and the number of diarrhoeal stools per day (ρ = 0.52, p < 0.001) as did CRP (ρ = 0.52, p < 0.001). CONCLUSIONS: We found no association between cessation of diarrhoea and metronidazole or lactoferrin concentrations. There was a relationship between metronidazole concentrations and markers of inflammation and stool frequency.

Factors associated with change in radiographic damage scores in gout: a prospective observational study.

BACKGROUND/AIMS: Radiographic damage is frequently observed in patients with longstanding gout. The aim of this prospective observational study was to determine factors associated with change in radiographic damage scores in gout. METHODS: People with gout and disease duration <10 years were recruited into this prospective observational study. At the baseline visit, structured assessment was undertaken in 290 participants including detailed clinical examination and plain radiographs (XR) of the hands and feet. Participants were invited to attend a further study visit with repeat XR 3 years after the baseline visit. XR were scored for erosion and joint space narrowing according to the gout-modified Sharp/van der Heijde XR damage score. RESULTS: Age, subcutaneous tophus count and tender joint count were independently associated with XR damage score at the baseline visit. Paired serial XR were available for 140 participants. In stepwise linear regression analysis, change in total damage score over 3 years was positively associated with change in subcutaneous tophus count and baseline XR damage score, and inversely associated with baseline subcutaneous tophus count (model R2=0.39, p<0.001). Change in subcutaneous tophus count contributed most to the change in erosion score (partial R2 change=0.31, p<0.001), and baseline XR damage score contributed most to the change in narrowing score (partial R2 change=0.31, p<0.001). CONCLUSIONS: Development of new subcutaneous tophi and baseline radiographic damage are associated with progressive joint damage scores in people with gout. These data provide further evidence that the tophus plays a central role in bone erosion in gout.

Regulation of TIMP-1 in Human Placenta and Fetal Membranes by lipopolysaccharide and demethylating agent 5-aza-2'-deoxycytidine.

BACKGROUND: An appropriate transcriptional profile in the placenta and fetal membranes is required for successful pregnancy; any variations may lead to inappropriate timing of birth. Epigenetic regulation through reversible modification of chromatin has emerged as a fundamental mechanism for the control of gene expression in a range of biological systems and can be modified by pharmacological intervention, thus providing novel therapeutic avenues. TIMP-1 is an endogenous inhibitor of MMPs, and hence is intimately involved in maintaining the integrity of the fetal membranes until labor. OBJECTIVE AND METHODS: To determine if TIMP-1 is regulated by DNA methylation in gestational tissues we employed an in vitro model in which gestational tissue explants were treated with demethylating agent 5-aza-2'-deoxycytidine (AZA) and lipopolysaccharide (LPS). RESULTS: Quantitative Real-Time PCR (qRT-PCR) revealed that TIMP-1 transcription was significantly increased by combined treatment of AZA and LPS, but not LPS alone, in villous, amnion and choriodecidua explants after 24 and 48 hrs, whilst western blotting showed protein production was stimulated after 24 hrs only. Upon interrogation of the TIMP-1 promoter using Sequenom EpiTyper MassARRAY, we discovered sex-specific differential methylation, in part explained by x-linked methylation in females. Increased TIMP-1 in the presence of LPS was potentiated by AZA treatment, signifying that a change in chromatin structure, but not in DNA methylation at the promoter region, is required for transcriptional activators to access the promoter region of TIMP-1. CONCLUSIONS: Collectively, these observations support a potential role for pharmacological agents that modify chromatin structure to be utilized in the therapeutic targeting of TIMP-1 to prevent premature rupture of the fetal membranes in an infectious setting.

Outcomes After Endovascular Aortic Intervention in Patients With Connective Tissue Disease.

Importance: Endovascular treatment is not recommended for aortic pathologies in patients with connective tissue diseases (CTDs) other than in redo operations and as bridging procedures in emergencies. However, recent developments in endovascular technology may challenge this dogma. Objective: To assess the midterm outcomes of endovascular aortic repair in patients with CTD. Design, Setting, and Participants: For this descriptive retrospective study, data on demographics, interventions, and short-term and midterm outcomes were collected from 18 aortic centers in Europe, Asia, North America, and New Zealand. Patients with CTD who had undergone endovascular aortic repair from 2005 to 2020 were included. Data were analyzed from December 2021 to November 2022. Exposure: All principal endovascular aortic repairs, including redo surgery and complex repairs of the aortic arch and visceral aorta. Main Outcomes and Measures: Short-term and midterm survival, rates of secondary procedures, and conversion to open repair. Results: In total, 171 patients were included: 142 with Marfan syndrome, 17 with Loeys-Dietz syndrome, and 12 with vascular Ehlers-Danlos syndrome (vEDS). Median (IQR) age was 49.9 years (37.9-59.0), and 107 patients (62.6%) were male. One hundred fifty-two (88.9%) were treated for aortic dissections and 19 (11.1%) for degenerative aneurysms. One hundred thirty-six patients (79.5%) had undergone open aortic surgery before the index endovascular repair. In 74 patients (43.3%), arch and/or visceral branches were included in the repair. Primary technical success was achieved in 168 patients (98.2%), and 30-day mortality was 2.9% (5 patients). Survival at 1 and 5 years was 96.2% and 80.6% for Marfan syndrome, 93.8% and 85.2% for Loeys-Dietz syndrome, and 75.0% and 43.8% for vEDS, respectively. After a median (IQR) follow-up of 4.7 years (1.9-9.2), 91 patients (53.2%) had undergone secondary procedures, of which 14 (8.2%) were open conversions. Conclusions and Relevance: This study found that endovascular aortic interventions, including redo procedures and complex repairs of the aortic arch and visceral aorta, in patients with CTD had a high rate of early technical success, low perioperative mortality, and a midterm survival rate comparable with reports of open aortic surgery in patients with CTD. The rate of secondary procedures was high, but few patients required conversion to open repair. Improvements in devices and techniques, as well as ongoing follow-up, may result in endovascular treatment for patients with CTD being included in guideline recommendations.

Population-specific factors associated with fractional excretion of uric acid.

BACKGROUND: Reduced renal clearance of uric acid is a major contributor to hyperuricemia. The aim of this study was to examine clinical and genetic variables associated with fractional excretion of uric acid (FEUA). METHODS: Participants (with and without gout) in the Genetics of Gout in Aotearoa study with available genotyping and FEUA data were included (n = 1713). Ten FEUA-associated loci detected within a genome-wide association study for serum urate in a European population were analysed. A polygenic score for FEUA was calculated in each ancestry group to model the cumulative effects of the genetic variants on FEUA. Associations between FEUA and both clinical variables and polygenic score were tested using linear regression models. RESULTS: The mean (SD) FEUA was 5.13 (2.70) % in Eastern Polynesian participants, 4.70 (5.89) % in Western Polynesian participants, and 5.89 (2.73) % in New Zealand European participants. Although association with FEUA was observed for SLC2A9 rs11942223 in New Zealand European participants (P = 2.39 × 10- 8), this association was not observed in Eastern or Western Polynesian participants. The polygenic score was positively associated with FEUA in all ancestry groups. In New Zealand European participants, body mass index, diuretic use, polygenic score, and male sex were associated with FEUA and explained 22% of FEUA variance in the regression model. In Eastern and Western Polynesian participants, the tested variables explained 10% and 4% of FEUA variance respectively. CONCLUSIONS: Both clinical and genetic variables contribute to renal clearance of uric acid. SLC2A9 exerts effects on FEUA variance in people of European ancestry, but not in those of Polynesian ancestry. There is a large unexplained variance in FEUA, particularly in people of Polynesian ancestry.

Development and validation of a liquid chromatography tandem mass spectrometry assay for the measurement of faecal metronidazole.

BACKGROUND: Metronidazole is an oral antibiotic which is widely used in the treatment of patients with Clostridium difficile associated disease. METHODS: This article describes the validation of a LC-MS/MS assay for the measurement of metronidazole in human faecal samples. RESULTS: Matrix matched and aqueous standards showed no significant difference in performance for the routine calibration of the assay. D4 deuterated metronidazole internal standard eluted with a different retention time to the undeuterated metronidazole on chromatography, hence zidovudine was used as an internal standard. Ion suppression was noted for both metronidazole and zidovudine due to unidentified compounds present in the faecal matrix and this was improved by extracting a smaller quantity of faeces and diluting the extract prior to analysis. Measurement uncertainty was 13% at 28,400ng/ml, 7.2% at 3300ng/ml, 3.9% at 320ng/ml, 13.6% at 109ng/ml and 30.9% at 20ng/ml. The assay was shown to be linear on dilution and the sensitivity of the assay was superior to HPLC assays using UV detection. The limit of detection was 5ng/ml, the limit of quantitation was 66ng/ml and the upper limit of the working range was 30,000ng/ml. Patient samples were stable at -20°C for 12months and extracted faecal samples were stable on storage for 1week at 4°C. There were no specific requirements for patient preparation or time of sample collection relative to taking metronidazole. CONCLUSIONS: Metronidazole can be quantified in faecal samples using LC-MS/MS which opens up opportunities for further research in this area.

Predictors of Mortality in People with Recent-onset Gout: A Prospective Observational Study.

OBJECTIVE: To determine mortality rates and predictors of death at baseline in people with a recent onset of gout. METHODS: People with gout disease duration < 10 years were recruited from primary and secondary care settings. Comprehensive clinical assessment was completed at baseline. Participants were prospectively followed for at least 1 year. Information about death was systematically collected from primary and secondary health records. Standardized mortality ratios (SMR) were calculated and risk factors for mortality were analyzed using Cox proportional hazard regression models. RESULTS: The mean (SD) followup duration was 5.1 (1.6) years (a total 1511 patient-yrs accrued). Of the 295 participants, 43 (14.6%) had died at the time of censorship (SMR 1.96, 95% CI 1.44-2.62). In the reduced Cox proportional hazards model, these factors were independently associated with an increased risk of death from all causes: older age (70-80 yrs: HR 9.96, 95% CI 3.30-30.03; 80-91 yrs: HR 9.39, 95% CI 2.68-32.89), Maori or Pacific ethnicity (HR 2.48, 95% CI 1.17-5.29), loop diuretic use (HR 3.99, 95% CI 2.15-7.40), serum creatinine (per 10 µmol/l change; HR 1.04, 95% CI 1.00-1.07), and the presence of subcutaneous tophi (HR 2.85, 95% CI 1.49-5.44). The presence of subcutaneous tophi was the only baseline variable independently associated with both cardiovascular (CV) cause of death (HR 3.13, 95% CI 1.38-7.10) and non-CV cause of death (HR 3.48, 95% CI 1.25-9.63). CONCLUSION: People with gout disease duration < 10 years have an increased risk of death. The presence of subcutaneous tophi at baseline is an independent predictor of mortality, from both CV and non-CV causes.

Illness Perceptions and Mortality in Patients With Gout: A Prospective Observational Study.

OBJECTIVE: To examine whether illness perceptions independently predict mortality in early-onset gout. METHODS: Between December 2006 and January 2014, a total of 295 participants with early-onset gout (<10 years) were recruited in Auckland and Wellington, New Zealand. The participants were followed up until February 2015, and mortality information was collected. Participants with complete data were included in the current study (n = 242). Cox proportional hazards models were used to examine the association between illness perceptions and mortality risk, after adjustment for covariates associated with disease severity and mortality in gout. RESULTS: In a Cox proportional hazards model adjusted for predictors of disease severity and mortality in gout (number of tophi, serum urate level, and frequency of flares), consequence beliefs, identity beliefs, concern beliefs, and emotional response to gout were associated with all-cause mortality (hazard ratios [HRs] 1.29, 1.15, 1.18, and 1.19, respectively; P < 0.05 for all). In the fully saturated model, the association between consequence beliefs and mortality remained robust after additional adjustment for ethnicity, disease duration, diuretic use, serum creatinine, and pain score (HR 1.18 [95% confidence interval 1.02-1.37]; P = 0.029). CONCLUSION: Negative beliefs about the impact of gout and severity of symptoms, as well as concerns about gout and the emotional response to gout, were independently associated with all-cause mortality. Illness perceptions are important and potentially modifiable risk factors to target in future interventions.

Faecal calprotectin as a marker for oesophago-gastric cancer.

BACKGROUND: Faecal calprotectin has been shown to be useful as a non-invasive screening test to differentiate functional from organic bowel disease, and it has been noted to be elevated in colorectal cancer. The aim of this study was to describe concentrations of faecal calprotectin in patients with oesophago-gastric cancer and investigate any potential discriminatory power of the test. PATIENTS: Faecal calprotectin was measured in samples from 39 patients with known oesophago-gastric cancer and in 191 samples from control subjects. RESULTS: The median calprotectin concentration was < 20 µg/g (range < 20-421 µg/g) in control subjects and 97 µg/g (range < 20-940 µg/g) in patients with oesophago-gastric cancer (P < 0.001). A receiver operating characteristic curve gave an area under the ROC curve of 0.84 and a sensitivity of 76.9% (95% CI: 63.7-90.1%) and specificity of 88.0% (95% CI: 83.3-92.6%) at a cutoff of 50 µg/g. CONCLUSION: Faecal calprotectin is elevated in patients with cancer of the upper gastrointestinal tract. This study suggests that calprotectin may be promising in discriminating cancer patients from controls, but further work is required to explore any potential role of faecal calprotectin in screening for, or diagnosis of, oesophago-gastric cancer.

Regulation of MT1-MMP/MMP-2/TIMP-2 axis in human placenta.

Matrix metalloproteinases (MMPs) and specific endogenous tissue inhibitors of metalloproteinases (TIMPs) mediate rupture of the fetal membranes in both physiological and pathological conditions. MMPs and TIMPs are subject to regulation by DNA methylation in human malignancies and pre-eclampsia. To determine if membrane type 1 MMP (MT1-MMP), MMP2, and TIMP2 are regulated by DNA methylation in human placentas, we employed an in vitro model where human placental tissues were collected at term gestation and cultured with methylation inhibiting agent 5-AZA-2'-deoxycytidine (AZA) and lipopolysaccharide. The results suggest that DNA methylation is not directly involved in the regulation of MT1-MMP in placental tissue; however, remodeling of chromatin by a pharmacologic agent such as AZA potentiates an infection-related increase in MT1-MMP. MT1-MMP is a powerful activator of MMP2 and this action, coupled with either no change or a decrease in TIMP2 concentrations, favors a gelatinolytic state leading to extracellular matrix degradation, which could predispose fetal membranes to rupture prematurely during inflammation.

Expression and regulation of DNA methyltransferases in human endometrium.

The messenger RNA of the DNA methyltransferases DNMT3a and DNMT3b are expressed temporally in the endometrium across the menstrual cycle, as is the steroid hormone regulation of DNMT1, DNMT3a, and DNMT3b. This suggests that DNA methylation in endometrium is changeable during the menstrual cycle and potentially alters gene expression.

Cytokine regulation during the formation of the fetal-maternal interface: focus on cell-cell adhesion and remodelling of the extra-cellular matrix.

The establishment of human pregnancy requires the orchestration of substantial cell differentiation and tissue remodelling processes in the context of a complex dialogue between the receptive endometrium and the implanting blastocyst, and is therefore dependent upon a complex sequence of signalling events. Cytokines play an important role in each step of implantation, modulating expression of adhesion molecules on both the fetal and maternal surfaces, regulating expression of the proteases that remodel the extra-cellular matrix, and promoting invasion and differentiation of trophoblasts. Here we review the role of cytokines in regulating the establishment of the fetal-maternal interface, with a particular focus on regulation of the functional expression of CAMs, the ECM and of the proteinases that modulate their function.