Induction of breast cancer cell apoptosis by novel thiouracil-fused heterocyclic compounds through boosting of Bax/Bcl-2 ratio and DFT study.

Breast cancer is a common public health disease causing mortality worldwide. Thus, providing novel chemotherapies that tackle breast cancer is of great interest. In this investigation, novel pyrido[2,3-d]pyrimidine derivatives 3,4,(6a-c),(8a,b),9-20 were synthesized and characterized using a variety of spectrum analyses. The geometric and thermal parameters of the novel thiouracil derivatives 3,4,6a,(8a,b),11,12,17,18, 19 were measured using density functional theory (DFT) via DFT/B3LYP/6-31 + G(d,p) basis set. All synthesized compounds were evaluated by MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) method using MCF-7 and MDA-MB-231 breast cancerous cells, compound 17 had the maximum anticancer activity against both breast cancerous cells, recording the lowest half-maximal inhibitory concentration (IC50) values (56.712 µg/mL for MCF-7 cells and 48.743 µg/mL for MDA-MB-231 cells). The results were confirmed in terms of the intrinsic mechanism of apoptosis, where compound 17 had the highest percentage in the case of both cancer cells and recorded Bax (Bcl-2 associated X)/Bcl-2 (B-cell lymphoma 2) ratio 17.5 and 96.667 for MCF-7 and MDA-MB-231 cells, while compound 19 came after 17 in the ability for induction of apoptosis, where the Bax/Bcl-2 ratio was 15.789 and 44.273 for both cancerous cells, respectively. Also, compound 11 recorded a high Bax/Bcl-2 ratio for both cells. The safety of the synthesized compounds was applied on normal WI-38 cells, showing minimum cytotoxic effect with undetectable IC50. Compounds 17, 11, and 19 recorded a significant increase of p53 upregulated modulator of apoptosis (PUMA) expression levels in the cancerous cells. The DFT method was also used to establish a connection between the experimentally determined values of the present investigated compounds and their predicted quantum chemical parameters. It was concluded that Compounds 17, 11, and 19 had anti-breast cancer potential through the induction of apoptotic Bax/Bcl-2 and PUMA expression levels.

Imidazo[1,2-b]pyridazines as inhibitors of DYRK kinases.

Selective inhibitors of DYRK1A are of interest for the treatment of cancer, Type 2 diabetes and neurological disorders. Optimization of imidazo [1,2-b]pyridazine fragment 1 through structure-activity relationship exploration and in silico drug design efforts led to the discovery of compound 17 as a potent cellular inhibitor of DYRK1A with selectivity over much of the kinome. The binding mode of compound 17 was elucidated with X-ray crystallography, facilitating the rational design of compound 29, an imidazo [1,2-b]pyridazine with improved kinase selectivity with respect to closely related CLK kinases.

Discovery of a stilbenoid-flavanone hybrid as an antitumor Wnt/ß-catenin signaling pathway inhibitor.

A series of designed stilbenoid-flavanone hybrids featuring sp3-hybridized C2 and C3 atoms of C-ring was evaluated against colorectal cancers presented compounds 4, 17, and 20 as the most potential compounds among explored compounds. Evaluation of the anticancer activity spectrum of compounds 4, 17, and 20 against diverse solid tumors presented compounds 17 and 20 with interesting anticancer spectrum. The potencies of compounds 17 and 20 were assessed in comparison with FDA-approved anticancer drugs. Compound 17 was the, in general, the most potent showing low micromolar GI50 values that were more potent than the standard FDA-approved drugs against several solid tumors including colon, brain, skin, renal, prostate and breast tumors. Compound 17 was subjected for evaluation against normal cell lines and was subjected to a mechanism study in HCT116 colon cancer cells which presented it as an inhibitor of Wnt signaling pathway triggering G2/M cell cycle arrest though activation of p53-p21 pathway as well as intrinsic and extrinsic apoptotic death of colon cancer cells. Compound 17 might be a candidate for further development against diverse solid tumors including colon cancer.

Evaluation of the technique of localization of the epidural space with the aid of a peripheral nerve stimulator and epidurographic, comparing two techniques for determining the infused volume, in rabbits (Oryctolagus Cuniculus).

Veterinary care for rabbits has been growing, and, consequently, the anesthetic and analgesic management of this species must be improved. The aim of the present study was to evaluate the technique of localization of the epidural space with the aid of a peripheral nerve stimulator and epidurographic, comparing two techniques for determining the infused volume in rabbits (Oryctolagus Cuniculus). In a prospective, randomized blinded study, six healthy New Zealand rabbits, adults, and weighing from 2.2 kg to 3.8 kg received two treatments, at 1 week intervals: 0.33 mL/kg (treatment I) or 0.05 mL per centimeter of the spine (treatment II) of ioexol epidurally. In both treatments, a peripheral nerve stimulator (2 Hz, 0.25 mA and 0.1 milliseconds) was used to determine the location of the epidural space. Latero-lateral and ventro-dorsal radiographs were taken after five (T5) and twenty-five minutes (T25) of iohexol administration. The epidural space was correctly accessed in 92% of attempts. Treatment I received a smaller volume of contrast than treatment II, 1.0 ± 0.2 mL versus 2.1 ± 0.1 mL (mean ± standard deviation), respectively (p = 0.007). At T5, the cranial progression of the contrast varied between L4 and L5 in treatment I, and L5 and T10 in treatment II. At T25, no contrast was observed in any rabbit. In conclusion, peripheral nerve stimulator aided in accessing the lumbosacral epidural space, and the administration of 0.05 mL per centimeter of the spine resulted in greater cranial progression of contrast.

Expression of GRP78 and its copartners in HEK293 and pancreatic cancer cell lines (BxPC-3/PANC-1) exposed to MRI and CT contrast agents.

Endoplasmic reticulum (ER) stress-associated chaperones trigger a defense mechanism called as unfolded protein response (UPR) which can manage apoptosis and be determinative in cell fate. Both anticancer drug effects and potential toxicity effects of magnetic resonance imaging (MRI) and computed tomography (CT) contrast agents were aimed to be evaluated. For this purpose, we investigated expression profiles of endoplasmic reticulum stress-associated chaperone molecules in human pancreatic tumor lines BxPC-3 and PANC-1 and control human embryonic kidney cells 293 (HEK293) induced with a variety of gadolinium and iohexol contrast agents. Protein expression levels of ER stress-associated chaperones (master regulator: GRP78/Bip and its copartners: Calnexin, Ero1, PDI, CHOP, IRE1α and PERK) were evaluated with Western blotting. Expression levels at mRNA level were also assessed for GRP78/Bip and CHOP with real-time PCR. Induction of cells was carried out with four different Gd-based contrast agents (GBCAs): (Dotarem, Optimark, Primovist and Gadovist) and two different iohexol agents (Omnipol, Omnipaque). CT contrast agents tested in the study did not result in significant ER stress in HEK293 cells. However, they do not seem to have theranostic potential in pancreas cancer through ER pathway. The potential efficiency of macrocyclic MRI contrast agents to provoke apoptosis via ER stress-associated chaperones in BxPC-3 cells lends credibility for their future theranostic use in pancreas cancer as long as undesired toxicity effects were carefully considered. ER stress markers and/or contrast agents seem to have promising potential to be translated into the clinical practice to manage pancreas cancer progression.

Comparison of Diatrizoate and Iohexol for Patient Acceptance and Fecal-Tagging Performance in Noncathartic CT Colonography.

OBJECTIVE: The aim of this study was to compare diatrizoate and iohexol regarding patient acceptance and fecal-tagging performance in noncathartic computed tomography colonography. METHODS: This study enrolled 284 volunteers with fecal tagging by either diatrizoate or iohexol at an iodine concentration of 13.33 mg/mL and an iodine load of 24 g. Patient acceptance was rated on a 4-point scale of gastrointestinal discomfort. Two gastrointestinal radiologists jointly analyzed image quality, fecal-tagging density and homogeneity, and residual contrast agent in the small intestine. The results were compared by the generalized estimating equation method. RESULTS: Patient acceptance was comparable between the 2 groups (3.95 ± 0.22 vs 3.96 ± 0.20, P = 0.777). The diatrizoate group had less residual fluid and stool than the iohexol group ( P = 0.019, P = 0.004, respectively). There was no significant difference in colorectal distention, residual fluid, and stool tagging quality between the 2 groups (all P 's > 0.05). The mean 2-dimensional image quality score was 4.59 ± 0.68 with diatrizoate and 3.60 ± 1.14 with iohexol ( P < 0.001). The attenuation of tagged feces was 581 ± 66 HU with diatrizoate and 1038 ± 117 HU with iohexol ( P < 0.001). Residual contrast agent in the small intestine was assessed at 55.3% and 62.3% for the diatrizoate group and iohexol group, respectively ( P = 0.003). CONCLUSIONS: Compared with iohexol, diatrizoate had better image quality, proper fecal-tagging density, and more homogeneous tagging along with comparable excellent patient acceptance, and might be more suitable for fecal tagging in noncathartic computed tomography colonography.

Optical clearing of human skin: Molecular modeling and in vivo OCT study.

The results of in vivo immersion optical clearing of human skin under the action of two different optical clearing agents (OCAs), such as an aqueous sucrose solution and a radiographic contrast agent Omnipaque™ 300 (iohexol), were obtained with the use of optical coherence tomography (OCT) method. The rate of reduction of light scattering coefficient, obtained through an averaged A-scan of the OCT image in the region of dermis within the depths from 350 to 700 µm, were determined to evaluate the efficiency of optical clearing (EOC). The correlations between the EOC and the energy of intermolecular interaction of OCAs with a fragment of collagen peptide have been established as a result of molecular modeling by quantum chemistry methods HF/STO3G/DFT/B3LYP/6-311G(d) of a number of OCAs (glycerol, iohexol, sucrose, ribose, fructose, glucose) with mimetic peptide of collagen (GPH)3 .

Predictors of augmented renal clearance based on iohexol plasma clearance in critically ill children.

BACKGROUND: Augmented renal clearance (ARC) holds a risk of subtherapeutic drug concentrations. Knowledge of patient-, disease-, and therapy-related factors associated with ARC would allow predicting which patients would benefit from intensified dosing regimens. This study aimed to identify ARC predictors and to describe ARC time-course in critically ill children, using iohexol plasma clearance (CLiohexol) to measure glomerular filtration rate (GFR). METHODS: This is a retrospective analysis of data from the "IOHEXOL" study which validated GFR estimating formulas (eGFR) against CLiohexol. Critically ill children with normal serum creatinine were included, and CLiohexol was performed as soon as possible after pediatric intensive care unit (PICU) admission (CLiohexol1) and repeated (CLiohexol2) after 48-72 h whenever possible. ARC was defined as CLiohexol exceeding normal GFR for age plus two standard deviations. RESULTS: Eighty-five patients were included; 57% were postoperative patients. Median CLiohexol1 was 122 mL/min/1.73 m2 (IQR 75-152). Forty patients (47%) expressed ARC on CLiohexol1. Major surgery other than cardiac surgery and eGFR were found as independent predictors of ARC. An eGFR cut-off value of 99 mL/min/1.73 m2 and 140 mL/min/1.73 m2 was suggested to identify ARC in children under and above 2 years, respectively. ARC showed a tendency to persist on CLiohexol2. CONCLUSIONS: Our findings raise PICU clinician awareness about increased risk for ARC after major surgery and in patients with eGFR above age-specific thresholds. This knowledge enables identification of patients with an ARC risk profile who would potentially benefit from a dose increase at initiation of treatment to avoid underexposure. TRIAL REGISTRATION: ClinicalTrials.gov NCT05179564, registered retrospectively on January 5, 2022.

Structure-Based Drug Design for Targeting IRE1: An in Silico Approach for Treatment of Cancer.

BACKGROUND: Endoplasmic Reticulum (ER) stress and Unfolded Protein Response (UPR) play a key role in cancer progression. The aggregation of incorrectly folded proteins in the ER generates ER stress, which in turn activates the UPR as an adaptive mechanism to fix ER proteostasis. Inositol-requiring enzyme 1 (IRE1) is the most evolutionary conserved ER stress sensor, which plays a pro-tumoral role in various cancers. Targeting its' active sites is one of the most practical approaches for the treatment of cancers. OBJECTIVE: In this study, we aimed to use the structure of 4µ8C as a template to produce newly designed compounds as IRE1 inhibitors. METHODS: Various functional groups were added to the 4µ8C, and their binding affinity to the target sites was assessed by conducting a covalent molecular docking study. The potential of the designed compound for further in vitro and in vivo studies was evaluated using ADMET analysis. RESULTS: Based on the obtained results, the addition of hydroxyl groups to 4µ8C enhanced the binding affinity of the designed compound to the target efficiently. Compound 17, which was constructed by the addition of one hydroxyl group to the structure of 4µ8C, can construct a strong covalent bond with Lys907. The outcomes of ADMET analysis indicated that compound 17 could be considered a drug-like molecule. CONCLUSION: Our results revealed that designed compound 17 could inhibit IRE1 activity. Therefore, this designed compound is a remarkable inhibitor of IRE1 and introduces a promising therapeutic strategy for cancer treatment.

In vitro characteristics of Epirubicin-loaded thermosensitive liquid embolic agent.

OBJECTIVE: To investigate the drug loading and release rate of epirubicin-loaded thermosensitive liquid embolic agents in vitro. MATERIALS AND METHODS: The drug loading and stability of epirubicin-loaded thermosensitive liquid embolic agents with or without iopromide were determined by high-performance liquid chromatography, and the same method was used to determine the drug release rate of thermosensitive liquid embolic agents at different time points. RESULTS: For epirubicin-loaded thermosensitive liquid embolic agents without iopromide, the average drug loading after filtration by membrane was (0.78 ± 0.02) mg and the drug loading rate was (16.1 ± 0.35)%, while the average drug loading without membrane was (0.73 ± 0.06) mg and the drug loading rate was (15.07 ± 1.17)%. After adding iopromide, the drug loading capacity was measured from 0 h-24 h solution and the drug loading was calculated indirectly and conclude that the drug loading capacity of thermosensitive liquid embolic agents decreased or disappeared. The sustained release rate of epirubicin from 0 to 48 hours was 42.65% in 48 hours. CONCLUSION: Epirubicin can be successfully loaded into the thermosensitive liquid embolic agents with good stability and sustained release. After adding iopromide, the drug loading capacity of thermosensitive liquid embolic agents decreased or disappeared.

Contrast media extravasation injury: a prospective observational cohort study.

OBJECTIVE: To identify the risk factors for moderate and severe contrast media extravasation and provide effective guidance to reduce the degree of extravasation injuries. METHODS: We observed 224 adult patients who underwent contrast media extravasation at Xiangya Hospital of Central South University, Hunan Provincial Maternal and Child Healthcare Hospital, and Xiangya Changde Hospital, Hunan Province between January 1, 2018 and December 31, 2022. Risk factors for moderate extravasation injuries were evaluated using univariate and multivariate logistic regression. RESULTS: Among 224 patients, 0 (0%) had severe, 18 (8.0%) had moderate, and 206 (92.0%) had mild contrast media extravasation injury. Multivariate logistic regression analysis revealed malignant tumors (odds ratio [OR] = 6.992, 95% confidence interval [CI]: 1.674-29.208), Iohexol (OR = 9.343, 95% CI 1.280-68.214), large-volume (> 50 mL) extravasation (OR = 5.773, 95% CI 1.350‒24.695), and injection site (back of the hand) (OR = 13.491, 95% CI 3.056-59.560) as independent risk factors for moderate injury. CONCLUSION: Risk factors for moderate contrast media extravasation injury are malignant tumors, iohexol, large-volume (> 50 mL) extravasation, and back-of-the-hand injection. Analysis of these risk factors can help reduce the degree of injury after extravasation. CLINICAL RELEVANCE STATEMENT: High-risk patients with extravasation support should choose the appropriate contrast media type, avoiding back-of-the-hand injections. We recommend that patients with cancer be implanted with a high-pressure resistant central venous catheter and receive effective measures to timely detect and reduce extravasation.

Contrast media-induced nephropathy in Tunisia: prospective case-control study with cardio-nephrological monitoring.

Introduction: vascular opacification using iodinated contrast media (ICM) is often the primary diagnostic and therapeutic approach. However, the risk of post-injection nephrotoxicity of ICM is significantly higher in patients with underlying nephropathy. This study aimed to determine the incidence of Contrast Media Induced Nephropathy (CMIN) and identify predictive factors for its occurrence in patients from a cardiology department. Methods: our prospective study involved 158 patients who underwent coronary angiography or angioplasty at the cardiology department between December 2017 and May 2018. Two types of ICM were used in our study: Iopromide and Iohexol. All patients received either physiological serum (9‰) or bicarbonate serum (14‰) intravenously for hydration. We defined impaired renal function as an increase in creatinine ranging from 10 to 26 µmol/L, while CMIN was defined as an increase in serum creatinine exceeding 26.5 µmol/L. We investigated the factors associated with CMIN using logistic regression analysis. Results: the mean age of our patients was 60 ± 11 years (range: 29-82), with a predominance of men 63.9% (n=101). The most common cardiovascular risk factors were tobacco (36.1%, n = 57), diabetes (48.1%, n =76), hypertension (55%, n = 87). Pre-procedural creatinine averaged 81.1 ± 47.3 µmol / L with extremes ranging from 39 to 600 µmol / L. The median Mehran risk score was 3.2 (range: 0- 15). The interventional cardiology act consisted of coronary angiography in 86.2% (n=136) of cases, coronary angioplasty in 2.5% (n=4) of cases. We used iohexol and iopromide in 57.6% (n=91) and 42.4% (n=67) of cases, respectively. The overall incidence of CMIN was 9.5% (n=9). The multivariable regression analysis identified 4 risk factors independently linked to the occurrence of CMIN which were Pre-existing renal failure (OR: 6.05, 95%CI [1.23-29.62], p = 0.026), anemia (OR: 0.043, CI [1.03-8.96], p = 0.043), the toxic dose of PC (OR: 4.7, CI [1.28-17.7], p=0.02), and at a Mehran score = 11 (OR: 3.7, CI [0.88-15.6], p=0.036). Conclusion: the most effective approach for CMIN is prevention, which focuses on addressing modifiable risk factors to minimize the risk especially in patients with pre-existing renal failure.

Ambulatory blood pressure as risk factor for long-term kidney function decline in the general population: a distributional regression approach.

The results of randomized controlled trials are unclear about the long-term effect of blood pressure (BP) on kidney function assessed as the glomerular filtration rate (GFR) in persons without chronic kidney disease or diabetes. The limited duration of follow-up and use of imprecise methods for assessing BP and GFR are important reasons why this issue has not been settled. Since a long-term randomized trial is unlikely, we investigated the association between 24-h ambulatory BP (ABP) and measured GFR in a cohort study with a median follow-up of 11 years. The Renal Iohexol Clearance Survey (RENIS) cohort is a representative sample of persons aged 50 to 62 years without baseline cardiovascular disease, diabetes, or kidney disease from the general population of Tromsø in northern Norway. ABP was measured at baseline, and iohexol clearance at baseline and twice during follow-up. The study population comprised 1589 persons with 4127 GFR measurements. Baseline ABP or office BP components were not associated with the GFR change rate in multivariable adjusted conventional regression models. In generalized additive models for location, scale, and shape (GAMLSS), higher daytime systolic, diastolic, and mean arterial ABP were associated with a slight shift of the central part of the GFR distribution toward lower GFR and with higher probability of GFR < 60 mL/min/1.73 m2 during follow-up (p < 0.05). The use of a distributional regression method and precise methods for measuring exposure and outcome were necessary to detect an unfavorable association between BP and GFR in this study of the general population.

Incomplete Recovery from the Radiocontrast-Induced Dysregulated Cell Cycle, Adhesion, and Fibrogenesis in Renal Tubular Cells after Radiocontrast (Iohexol) Removal.

Contrast-induced nephropathy (CIN) is one of the most common causes of acute kidney injury (AKI). However, management is still limited, and the cellular response to radiocontrast removal for CIN remains unclear. This study aimed to explore the latent effects of iohexol in cultured renal tubular cells with or without the removal of iohexol by medium replacement. HK2 renal tubular cells were subcultured 24 h before use in CIN experiments. Three treatment groups were established: the control, a radiocontrast (iohexol)-only group at 75 mg I/mL (I-75), and iohexol exposure for 24 h with culture medium replacement (I-75/M). Cell cycle arrest, fibrogenic mediator assays, cell viability, cell function, and cell-cycle-related protein expression were compared between groups. Iohexol induced numerous changes in HK2 renal tubular cells, such as enlarged cell shape, cell cycle arrest, increased apoptosis, and polyploidy. Iohexol inhibited the expression of cyclins, CDKs, ZO-1, and E-cadherin but conversely enhanced the expression of p21 and fibrosis-related genes, including TGF-ß1, CTGF, collagen I, collagen III, and HIF-1α within 60 hr after the exposure. Except for the recovery from cell cycle arrest and cell cycle gene expression, notably, the removal of iohexol by medium replacement could not fully recover the renal tubular cells from the formation of polyploid cells, the adhesion or spreading, or the expression of fibrosis-related genes. The present study demonstrates, for the first time, that iohexol exerts latent cytotoxic effects on cultured renal tubular cells after its removal, suggesting that these irreversible cell changes may cause the insufficiency of radiocontrast reduction in CIN, which is worth investigating further.

Iodinated non-ionic contrast medium does not inhibit Escherichia coli growth in ex vivo inoculated canine urine.

OBJECTIVE: To investigate the effect of iohexol on standardized quantitative urine culture results in dogs. The authors hypothesized that the presence of iohexol in inoculated urine samples would result in lower bacterial concentrations (CFU/mL) and, therefore, decrease culture sensitivity. SAMPLE: Urine samples were aseptically collected during cystoscopy from a single client-owned dog untreated with antimicrobials. PROCEDURES: An experimental controlled study. The urine sample was divided into 38 aliquots (0.5 mL each) that were used as negative controls or inoculated with an equal amount of Escherichia coli (105 CFU/mL). Different volumes (0.1 and 0.5 mL) of contrast or saline were added to the aliquots and quantitative culture results were compared. Two different incubation times between the preparation of aliquots and culture were evaluated (15 minutes and 24 hours). RESULTS: All aliquots from samples inoculated with E. coli (positive controls and iohexol-group) had the same reported quantitative result (104 CFU/mL). No growth was reported for the negative controls. Iohexol did not show any anti-E. coli properties in canine urine cultures for dilutions up to 1:2 contrast:urine and concentrations up to 120 mgI/mL. No difference was reported when iohexol was incubated with inoculated urine for 15 minutes or 24 hours. CLINICAL RELEVANCE: Based on the experimental in vitro conditions described, administration of iohexol before the collection of urine during urologic procedures does not negatively impact the isolation and growth of E. coli.

Comparison of Cystatin C and Creatinine in the Assessment of Measured Kidney Function during Critical Illness.

BACKGROUND: Incomplete recovery of kidney function is an important adverse outcome in survivors of critical illness. However, unlike eGFR creatinine, eGFR cystatin C is not confounded by muscle loss and may improve identification of persistent kidney dysfunction. METHODS: To assess kidney function during prolonged critical illness, we enrolled 38 mechanically ventilated patients with an expected length of stay of >72 hours near admission to intensive care unit (ICU) in a single academic medical center. We assessed sequential kidney function using creatinine, cystatin C, and iohexol clearance measurements. The primary outcome was difference between eGFR creatinine and eGFR cystatin C at ICU discharge using Bayesian regression modeling. We simultaneously measured muscle mass by ultrasound of the rectus femoris to assess the confounding effect on serum creatinine generation. RESULTS: Longer length of ICU stay was associated with greater difference between eGFR creatinine and eGFR cystatin C at a predicted rate of 2 ml/min per 1.73 m 2 per day (95% confidence interval [CI], 1 to 2). By ICU discharge, the posterior mean difference between creatinine and cystatin C eGFR was 33 ml/min per 1.73 m 2 (95% credible interval [CrI], 24 to 42). In 27 patients with iohexol clearance measured close to ICU discharge, eGFR creatinine was on average two-fold greater than the iohexol gold standard, and posterior mean difference was 59 ml/min per 1.73 m 2 (95% CrI, 49 to 69). The posterior mean for eGFR cystatin C suggested a 22 ml/min per 1.73 m 2 (95% CrI, 13 to 31) overestimation of measured GFR. Each day in ICU resulted in a predicted 2% (95% CI, 1% to 3%) decrease in muscle area. Change in creatinine-to-cystatin C ratio showed good longitudinal, repeated measures correlation with muscle loss, R =0.61 (95% CI, 0.50 to 0.72). CONCLUSIONS: eGFR creatinine systematically overestimated kidney function after prolonged critical illness. Cystatin C better estimated true kidney function because it seemed unaffected by the muscle loss from prolonged critical illness. CLINICAL TRIAL REGISTRY NAME AND REGISTRATION NUMBER: Skeletal Muscle Wasting and Renal Dysfunction After Critical Illness Trauma - Outcomes Study (KRATOS), NCT03736005 .

Ox-LDL aggravates contrast-induced injury of renal tubular epithelial cells.

Hypercholesterolemia can aggravate contrast-induced acute kidney injury, and the exacerbation of renal tubular epithelial cell (RTEC) injury is a major cause. However, the exact mechanisms remain obscure. Mitophagy, a type of autophagy, selectively eliminates damaged mitochondria and reduces mitochondrial oxidative stress, which is strongly implicated in cell homeostasis and acute kidney injury. Oxidized low-density lipoprotein (Ox-LDL) is accumulated in hypercholesterolemia and has a cytotoxic effect. This study aimed to determine whether and how ox-LDL exacerbates contrast-induced injury in RTECs and to further explore whether PINK1/Parkin-dependent mitophagy is involved in this process. Iohexol and ox-LDL were used alone or in combination to treat HK-2 cells. Rapamycin pretreatment was utilized to enhance mitophagy. Cell viability, apoptosis, mitochondrial membrane potential (MMP) and mitochondrial reactive oxygen species (mtROS) were detected by cell counting kit-8, TUNEL staining, JC-1 kit and MitoSOX fluorescence, respectively. The expression of mitophagy-related proteins (including PINK1, Parkin, and so on) and cleaved caspase-3 was confirmed by western blot. Colocalization of MitoTracker-labeled mitochondria and LysoTracker-labeled lysosomes was observed by fluorescence microscopy to evaluate mitophagy. The results of our study showed that ox-LDL aggravated MMP decline, mtROS release and apoptosis in iohexol-treated HK-2 cells, accompanied by a further increased autophagy level. Enhancement of PINK1/Parkin-dependent mitophagy by rapamycin alleviated apoptosis and mitochondrial injury in HK-2 cells in response to iohexol under ox-LDL condition. Therefore, our findings indicate that ox-LDL aggravates contrast-induced injury of RTECs by increasing mitochondrial damage and mitochondrial oxidative stress, which may be associated with the relative insufficiency of PINK1/Parkin-dependent mitophagy.

Validation of eGFR for Detecting Associations Between Serum Protein Biomarkers and Subsequent GFR Decline.

SIGNIFICANCE STATEMENT: eGFR from creatinine, cystatin C, or both has been primarily used in search of biomarkers for GFR decline. Whether the relationships between biomarkers and eGFR decline are similar to associations with measured GFR (mGFR) decline has not been investigated. This study revealed that some biomarkers showed statistically significant different associations with eGFR decline compared with mGFR decline, particularly for eGFR from cystatin C. The findings indicate that non-GFR-related factors, such as age, sex, and body mass index, influence the relationship between biomarkers and eGFR decline. Therefore, the results of biomarker studies using eGFR, particularly eGFRcys, should be interpreted with caution and perhaps validated with mGFR. BACKGROUND: Several serum protein biomarkers have been proposed as risk factors for GFR decline using eGFR from creatinine or cystatin C. We investigated whether eGFR can be used as a surrogate end point for measured GFR (mGFR) when searching for biomarkers associated with GFR decline. METHODS: In the Renal Iohexol Clearance Survey, GFR was measured with plasma iohexol clearance in 1627 individuals without diabetes, kidney, or cardiovascular disease at baseline. After 11 years of follow-up, 1409 participants had one or more follow-up GFR measurements. Using logistic regression and interval-censored Cox regression, we analyzed the association between baseline levels of 12 serum protein biomarkers with the risk of accelerated GFR decline and incident CKD for both mGFR and eGFR. RESULTS: Several biomarkers exhibited different associations with eGFR decline compared with their association with mGFR decline. More biomarkers showed different associations with eGFRcys decline than with eGFRcre decline. Most of the different associations of eGFR decline versus mGFR decline remained statistically significant after adjustment for age, sex, and body mass index, but several were attenuated and not significant after adjusting for the corresponding baseline mGFR or eGFR. CONCLUSIONS: In studies of some serum protein biomarkers, eGFR decline may not be an appropriate surrogate outcome for mGFR decline. Although the differences from mGFR decline are attenuated by adjustment for confounding factors in most cases, some persist. Therefore, proposed biomarkers from studies using eGFR should preferably be validated with mGFR.

Accuracy of GFR Estimating Equations in Patients with Discordances between Creatinine and Cystatin C-Based Estimations.

SIGNIFICANCE STATEMENT: Large discordances between eGFR on the basis of creatinine (eGFR cr ) or cystatin C (eGFR cys ) are common in clinical practice. However, which GFR estimating equation (eGFR cr , eGFR cys , or eGFR cr-cys ) is most accurate in these settings is not known. In this real-world study of 9404 concurrent measurements of creatinine, cystatin C, and iohexol clearance, all three equations performed similarly when eGFR cr and eGFR cys were similar (45% of cases). However, with large discordances (55% of cases), eGFR cr-cys was much more accurate than either alone. These findings were consistent among individuals with cardiovascular disease, heart failure, diabetes mellitus, liver disease, and cancer who have been underrepresented in research cohorts. Thus, when eGFR cr and eGFR cys are largely discordant in clinical practice, eGFR cr-cys is more accurate than eGFR cr or eGFR cys . BACKGROUND: Cystatin C is recommended as a confirmatory test to eGFR when more precise estimates are needed for clinical decision making. Although eGFR on the basis of both creatinine and cystatin (eGFR cr-cys ) is the most accurate estimate in research studies, it is uncertain whether this is true in real-world settings, particularly when there are large discordances between eGFR based on creatinine (eGFR cr ) and that based on cystatin C (eGFR cys ). METHODS: We included 6185 adults referred for measured GFR (mGFR) using plasma clearance of iohexol in Stockholm, Sweden, who had 9404 concurrent measurements of creatinine, cystatin C, and iohexol clearance. The performance of eGFR cr , eGFR cys , and eGFR cr-cys was assessed against mGFR with median bias, P30 , and correct classification of GFR categories. We stratified analyses within three categories: eGFR cys at least 20% lower than eGFR cr (eGFR cys eGFR cr ). RESULTS: eGFR cr and eGFR cys were similar in 4226 (45%) samples, and among these samples all three estimating equations performed similarly. By contrast, eGFR cr-cys was much more accurate in cases of discordance. For example, when eGFR cys eGFR cr (8% of samples), the median biases were -4.5, 8.4, and 1.4 ml/min per 1.73m 2 . The findings were consistent among individuals with cardiovascular disease, heart failure, diabetes mellitus, liver disease, and cancer. CONCLUSIONS: When eGFR cr and eGFR cys are highly discordant in clinical practice, eGFR cr-cys is more accurate than either eGFR cr or eGFR cys .

Proenkephalin A as a marker for glomerular filtration rate in critically ill children: validation against gold standard iohexol GFR measurements.

OBJECTIVES: Accurate determination of glomerular filtration rate (GFR) is important. Several endogenous biomarkers exist for estimating GFR, yet, they have limited accuracy, especially in the paediatric population. Proenkephalin A 119-159 (PENK) is a novel and promising GFR marker, but its relation with age in children remains unknown. Also, the value of PENK has never been validated against measured GFR (mGFR) in children when compared to traditional GFR markers including serum creatinine (SCr), SCr-based estimated GFR (eGFR) and cystatin C (cysC). METHODS: Critically ill children and term-born neonates were included in this single-centre, prospective study. Iohexol-based mGFR, SCr, and cysC were determined in each patient. eGFR was calculated using the bedside Schwartz equation, incorporating SCr and height. Spearman correlation coefficients were calculated to determine the correlation between mGFR and PENK, SCr, cysC and eGFR. RESULTS: For 97 patients (56 children and 41 neonates), mGFR, SCr, cysC and PENK levels were available. PENK levels were higher in young children and decreased to adult PENK reference values around two years of age. PENK levels were highly correlated with mGFR (ρ=-0.88, p<0.001), and similar to mGFR-eGFR correlation (ρ=-0.87, p<0.001). For cysC and SCr the correlation with mGFR was lower (ρ=-0.77 and ρ=-0.46, respectively. Both p<0.001). CONCLUSIONS: The correlation of PENK with mGFR was as good as SCr-based eGFR-mGFR correlation. To determine the added value of PENK in paediatric clinical care and prior to implementation, PENK reference values are needed and the development and validation of a paediatric PENK-based eGFR equation is necessary.