The development of Bretschneider's histidine-tryptophan-ketoglutarate (HTK) cardioplegia solution represented a major advancement in cardiac surgery, offering significant myocardial protection. However, metabolic changes induced by this additive in the whole body have not been systematically investigated. Using an untargeted mass spectrometry-based method to deeply explore the urine metabolome, we sought to provide a holistic and systematic view of metabolic perturbations occurred in patients receiving HTK. Prospective urine samples were collected from 100 patients who had undergone cardiac surgery, and metabolomic changes were profiled using a high-performance chemical isotope labeling liquid chromatography-mass spectrometry (LC-MS) method. A total of 14,642 peak pairs or metabolites were quantified using differential 13C-/12C-dansyl labeling LC-MS, which targets the amine/phenol submetabolome from urine specimens. We identified 223 metabolites that showed significant concentration change (fold change > 5) and assembled several potential metabolic pathway maps derived from these dysregulated metabolites. Our data indicated upregulated histidine metabolism with subsequently increased glutamine/glutamate metabolism, altered purine and pyrimidine metabolism, and enhanced vitamin B6 metabolism in patients receiving HTK. Our findings provide solid evidence that HTK solution causes significant perturbations in several metabolic pathways and establish a basis for further study of key mechanisms underlying its organ-protective or potential harmful effects.

Patients with pneumonia and parapneumonic effusion (PPE) have elevated mortality and a poor prognosis. The aim of this study was to discover novel biomarkers to help distinguish between uncomplicated PPE (UPPE) and complicated PPE (CPPE). Using an iTRAQ-based quantitative proteomics, we identified 766 proteins in pleural effusions from PPE patients. In total, 45 of these proteins were quantified as upregulated proteins in CPPE. Four novel upregulated candidates (BPI, NGAL, AZU1, and calprotectin) were selected and further verified using enzyme-linked immunosorbent assays (ELISAs) on 220 patients with pleural effusions due to different causes. The pleural fluid levels of BPI, NGAL, AZU1, and calprotectin were significantly elevated in patients with CPPE. Among these four biomarkers, BPI had the best diagnostic value for CPPE, with an AUC value of 0.966, a sensitivity of 97%, and a specificity of 91.4%. A logistic regression analysis demonstrated a strong association between BPI levels > 10 ng/ml and CPPE (odds ratio = 341.3). Furthermore, the combination of pleural fluid BPI levels with LDH levels improved the sensitivity and specificity to 100% and 91.4%, respectively. Thus, our findings provided a comprehensive effusion proteome data set for PPE biomarker discovery and revealed novel biomarkers for the diagnosis of CPPE.

The role of fibroblasts in tissue fibrosis has been extensively studied. Activated fibroblasts, namely myofibroblasts, produce pathological extracellular matrix. CD248, a type I transmembrane glycoprotein, is expressed in fibroblasts after birth. In human chronic kidney disease, upregulated CD248 in myofibroblasts is linked to poor renal survival. In this study, we demonstrated a novel interaction between CD248 and macrophages to be a key step in mediating tissue fibrosis. CD248 was upregulated in myofibroblasts in murine models of renal and peritoneal fibrosis. Cd248 knockout (Cd248-/-) could attenuate both renal and peritoneal fibrosis. By parabiosis of GFP reporter mice and Cd248-/- mice, we showed that attenuation of renal fibrosis was associated with a decrease of macrophage infiltration in Cd248-/- mice. Moreover, decrease of chemokine (C-C motif) ligand 17 and Ccl22 was found in macrophages isolated from the fibrotic kidneys of Cd248-/- mice. Because galectin-3-deficient macrophages showed decreased Ccl17 and Ccl22 in fibrotic kidneys, we further demonstrated that CD248 interacted specifically with galectin-3 of macrophages who then expressed CCL17 to activate collagen production in myofibroblasts. Mice with DNA vaccination targeting CD248 showed decreased fibrosis. We thus propose that CD248 targeting should be studied in the clinical tissue fibrosis setting.

Esophageal squamous neoplasm presents a spectrum of different diatheses. A precise assessment for individualized treatment depends on the accuracy of the initial diagnosis. Detection relies on comprehensive and accurate white-light, iodine staining, and narrow-band imaging endoscopy. These methods have limitations in addition to its invasive nature and the potential risks related to the method. These limitations include difficulties in precise tumor delineation to enable complete resection, inflammation and malignancy differentiation, and stage determination. The resolution of these problems depends on the surgeon's ability and experience with available technology for visualization and resection. We proposed a method for identifying early esophageal cancerous lesion by endoscopy and hyperspectral endoscopic imaging. Experimental result shows the characteristic spectrum of a normal esophagus, precancerous lesion, canceration, and intraepithelial papillary capillary loop can be identified through principal component score chart. The narrow-band imaging (NBI) image shows remarkable spectral characteristic distribution, and the sensitivity and specificity of the proposed method are higher than those of other methods by ~0.8 and ~0.88, respectively. The proposed method enables the accurate visualization of target organs, it may be useful to capsule endoscope and telemedicine, which requires highly precise images for diagnosis.

Previous studies have shown that sirtuin 1 (Sirt1) is renoprotective; however, details regarding its distribution and functions in the kidney remain unknown. Here, we demonstrated that Sirt1 was mainly expressed in the tubulointerstitial cells of normal rat kidneys and was co-localized with aquaporin 2, indicating it may be involved in water/salt regulation. Renal Sirt1 expression increased in the non-glomerular cytoplasmic portion of the kidney after a 24-h fast, but no significant changes in Sirt1 expression occurred after water loading (50 mL/kg) or 24-h water deprivation. After consuming a low-salt (0.075%) or 60% calorie restriction diet for 7 days, Sirt1 expression in the rat kidney was significantly increased, whereas a high-salt (8%) diet did not change the level of Sirt1 expression. The low-salt diet also increased Sirt1 expression in the heart, muscle, brain, and fat tissues. The increased Sirt1 that was observed in rats on a low-salt diet was associated with increased ghrelin expression in the distal nephron, with both molecules exhibiting similar distribution patterns. An in vitro experiment suggested that ghrelin increases Sirt1 expression in cortical collecting duct cells by activating ghrelin receptors. Our study indicates that this 'ghrelin-Sirt1 system' may participate in regulating sodium reabsorption in the distal nephron.

Patients with complicated parapneumonic effusion (CPPE)/empyema have high morbidity and mortality, particularly when adequate management is delayed. We aimed to investigate novel dysregulated cytokines that can be used as biomarkers for infectious pleural effusions, especially for CPPE/empyema. Expression of 40 cytokines in parapneumonic effusions (PPE) was screened in the discovery phase, involving 63 patients, using a multiplex immunobead-based assay. Six cytokines were subsequently validated by enzyme-linked immunosorbent assays (ELISAs). We then used ELISA to further evaluate the diagnostic values and cutoff values of these cytokines as potential biomarkers in an expanded group that included 200 patients with uncomplicated parapneumonic effusion (UPPE), CPPE, empyema, transudates, other exudates, and malignant pleural effusion (MPE). The pleural levels of four cytokines (MIF, MIP-3α, IL-1β, ENA-78) were highest and significantly increased in CPPE/empyema compared with those in other etiologies. According to receiver operating characteristic curve analysis, the four cytokines (MIF, MIP-3α, IL-1β, and ENA-78) had areas under the curve (AUCs) greater than 0.710 for discriminating parapneumonic pleural effusion from noninfectious pleural effusions. In a comparison of nonpurulent CPPE with UPPE, logistic regression analysis revealed that pleural fluid MIF ≥ 12 ng/ml and MIP-3α ≥ 4.3 ng/ml had the best diagnostic value; MIF also displayed the highest odds ratio of 663 for nonpurulent CPPE, with 97.5% specificity, 94.44% sensitivity, and an AUC of 0.950. In conclusion, our results show that elevated MIF and MIP-3α may be used as novel biomarkers for PPE diagnosis, particularly in patients with CPPE/empyema; the findings indicate that dysregulated cytokine expression may provide clues about the pathogenesis of pleural infection.

Inflammatory bowel disease is a chronic and progressive inflammatory intestinal disease that includes two major types, namely ulcerative colitis and Crohn's disease (CD). CD is characterized by intestinal epithelial hyperplasia and inflammatory cell infiltration. Transfer of CD25-CD45RBhiCD4+ (naïve) T cells into immunodeficiency mice induces autoimmune colitis with pathological lesions similar to CD and loss of body weight 4 weeks after cell transfer. However, weight loss neither has sufficient sensitivity nor totally matches the pathological findings of CD. To establish an early and sensitive indicator of autoimmune colitis model, the transferred T cell-induced colitis mouse model was modified by transferring luciferase-expressing donor T cells and determining the colitis by in vivo imaging system (IVIS). Colitis was detected with IVIS 7-10 days before the onset of body weight loss and diarrhea. IVIS was also applied in the dexamethasone treatment trial, and was a more sensitive indicator than body weight changes. All IVIS signals were parallel to the pathological abnormalities of the gut and immunological analysis results. In summary, IVIS provides both sensitive and objective means to monitor the disease course of transferred T cell-induced CD and fulfills the 3Rs principle of humane care of laboratory animals.

Activation of sirtuin 1 (Sirt1) attenuates unilateral ureteral obstruction (UUO)-induced inflammation and fibrosis, suggesting that Sirt1 may prevent tubulointerstitial fibrosis. In this study, we explored changes in the expression of Sirt1 in the kidneys of UUO-treated rats and evaluated the effects of Sirt1 activation or inhibition on renal pathology and mediators of UUO pathogenesis, especially angiotensin II and nuclear factor (NF)-κB, in rats and rat renal fibroblasts. Sirt1 expression increased in the obstructed kidney but not in the contralateral kidney and was mainly detected in tubulointerstitial cells. Resveratrol, a Sirt1 activator, decreased UUO-induced inflammation and fibrosis, while sirtinol, a Sirt1 inhibitor, enhanced UUO-induced inflammation. UUO increased renal angiotensin type 1 receptor (AT1R), NF-κB, monocyte chemotactic protein 1 (MCP-1), and fibronectin expression. Resveratrol attenuated these UUO-induced changes, whereas sirtinol enhanced them, with the exception of fibronectin. In renal fibroblasts, Sirt1 overexpression reduced AT1R and NF-κB levels, while Sirt1 knockdown had the opposite effects. Sirtinol increased the levels of AT1R, NF-κB, MCP-1, and connective tissue growth factor (CTGF), while resveratrol reduced AT1R levels. Our results suggested that Sirt1 inhibited AT1R and NF-κB expression in renal fibroblasts and that these mechanisms may play roles in alleviating UUO-induced damages.

Chronic ketamine abuse is associated with bladder dysfunction and cystitis. However, the effects of ketamine abuse on the urinary proteome profile and the correlations among urinary proteins, urinary ketamine (and metabolites) and clinicopathological features of ketamine-induced bladder dysfunction remain to be established. Here, we recruited 56 ketamine abusers (KA) and 40 age-matched healthy controls (HC) and applied the iTRAQ-based proteomics approach to unravel quantitative changes in the urine proteome profile between the two groups. Many of the differentially regulated proteins are involved in the complement and coagulation cascades and/or fibrotic disease. Among them, a significant increase in APOA1 levels in KA relative to control samples (392.1 ± 59.9 ng/ml vs. 13.7 ± 32.6 ng/ml, p < 0.0001) was detected via ELISA. Moreover, urinary ketamine, norketamine and dehydronorketamine contents (measured via LC-SRM-MS) were found to be positively correlated with overactive bladder syndrome score (OABSS) and APOA1 levels with urinary RBC, WBC, OABSS and numeric pain rating scale in KA. Collectively, our results may aid in developing new molecular tool(s) for management of ketamine-induced bladder dysfunction. Moreover, information regarding the differentially regulated proteins in urine of KA provides valuable clues to establish the molecular mechanisms underlying ketamine-induced cystitis.

Patients with kidney failure rely on life-saving peritoneal dialysis to facilitate waste exchange and maintain homeostasis of physical conditions. However, peritoneal dialysis often results in peritoneal fibrosis and organ adhesion that subsequently compromise the efficiency of peritoneal dialysis and normal functions of visceral organs. Despite rodent models provide clues on the pathogenesis of peritoneal fibrosis, no current large animal model which shares high degree of physiological and anatomical similarities to human is available, limiting their applications on the evaluation of pre-clinical therapeutic efficacy. Here we established for the first time, hypochlorite-induced porcine model of peritoneal fibrosis in 5-week-old piglets. We showed that administration 15-30 mM hypochlorite, a dose- and time-dependent severity of peritoneal fibrosis characterized by mesothelium fragmentation, αSMA+ myofibroblasts accumulation, organ surface thickening and type I collagen deposition were observed. We also demonstrated in vitro using human mesothelial cells that hypochlorite-induced fibrosis was likely due to necrosis, but not programmed apoptosis; besides, overexpression of IL1β, CX3CL1 and TGFβ on the peritoneal mesothelium in current model was detected, similar to observations from peritoneal dialysis-induced peritoneal fibrosis in human patients and earlier reported mouse model. Moreover, our novel antemortem evaluation using laparoscopy provided instant feedback on the progression of organ fibrosis/adhesion which allows immediate adjustments on treatment protocols and strategies in alive individuals that can not and never be performed in other animal models.

Acute kidney injury (AKI) is an important risk factor for incident chronic kidney disease (CKD). Clinical studies disclose that ensuing CKD progresses after functional recovery from AKI, but the underlying mechanisms remain illusive. Using a murine model representing AKI-CKD continuum, we show angiotensin II type 1a (AT1a) receptor signaling as one of the underlying mechanisms. Male adult CD-1 mice presented severe AKI with 20% mortality within 2 weeks after right nephrectomy and left renal ischemia-reperfusion injury. Despite functional recovery, focal tubular atrophy, interstitial cell infiltration and fibrosis, upregulation of genes encoding angiotensinogen and AT1a receptor were shown in kidneys 4 weeks after AKI. Thereafter mice manifested increase of blood pressure, albuminuria and azotemia progressively. Drinking water with or without losartan or hydralazine was administered to mice from 4 weeks after AKI. Increase of mortality, blood pressure, albuminuria, azotemia and kidney fibrosis was noted in mice with vehicle administration during the 5-month experimental period. On the contrary, these parameters in mice with losartan administration were reduced to the levels shown in control group. Hydralazine did not provide similar beneficial effect though blood pressure was controlled. These findings demonstrate that losartan can reduce ensuing CKD and mortality after functional recovery from AKI.

Higher baseline glomerular filtration rate (GFR) may yield subsequent steeper GFR decline, especially in patients with diabetes mellitus (DM). However, this correlation in patients with chronic kidney disease (CKD) and the presence or absence of DM remains controversial. We conducted a longitudinal cohort study in a single medical center between 2011 and 2018. Participants with CKD stage 1 to 3A were enrolled and divided into DM groups and non-DM groups, and then followed up at least every 6 months. We used a linear mixed regression model with centering time variable to overcome the problem of mathematical coupling in the analysis of the relation between baseline GFR and the changes, and compared the results from correct and incorrect specifications of the mixed models. A total number of 1002 patients with 285 diabetic and 717 non-diabetic persons was identified. The linear mixed regression model revealed a significantly negative correlation between baseline GFR and subsequent GFR change rate in both diabetic group and non-diabetic group (r =  - 0.44 [95% confidence interval [CI], - 0.69 to - 0.09]), but no statistical significance in non-diabetic group after within-subject mean centering of time variable (r =  - 0.09 [95% CI, - 0.41 to 0.25]). Our study showed that higher baseline GFR was associated with a subsequent steeper GFR decline in the DM group but not in the non-DM group among patients with early-stage CKD. Exact model specifications should be described in detail to prevent from a spurious conclusion.

Vascular development is regulated by complicated signals and molecules in vertebrates. In this study, we characterized a novel function of carboxypeptidase N1 (Cpn1) in the vasculature. We show that cpn1 mRNA is expressed in developing vessels. The knockdown of cpn1 by morpholino injection impairs the growth of intersegmental vessels (ISV) and caudal vein plexus (CVP), suggesting the role of cpn1 in vascular development. We showed that vascular defects are not caused by cell death but are due to the impairment of migration and proliferation. Consistent with vascular growth defects, loss of cpn1 affects the expression of the vascular markers flt4, mrc1, flk, stabilin, and ephrinb2. Furthermore, the overexpression of cpn1 impaired the growth of ISV and CVP, but the remodeling expression of vascular markers was different from the knockdown of cpn1, indicating the differential regulation mechanisms in cpn1-overexpressing embryos. We examine the interaction between cpn1 and multiple signals and observed that cpn1 is regulated by Notch/VEGF signals for ISV growth and likely regulates BMP signals for CVP patterning. In conclusion, we demonstrate that cpn1 has a critical role in the vascular development of zebrafish. We also reveal a fine-tune regulation of cpn1 that controls vascular patterning mediated by multiple signals.

The gene of protein interacting with C kinase 1 alpha (PICK1) has been implicated in schizophrenia, nevertheless, conflicting results existed. However, its role in cognitive function remains unclear. Besides, cognitive deficits impair the long-term outcome. We explored whether the polymorphisms of PICK1 (rs2076369, rs3952) affected cognitive functions in schizophrenic patients. We analyzed 302 patients and tested the differences of cognitive functions, clinical symptoms between genetic groups. We also used general linear model to analyze the effect of PICK1 genetic polymorphisms on cognitive functions. After adjustment for gender, age, education, the patients with rs2076369 G/T genotype showed better performance than T/T homozygotes in the summary score, global composite score, neurocognitive composite score, category fluency subtest, WAIS-III-Digit Symbol Coding subtest, working memory, WMS-III-Spatial Span (backward) subtest, MSCEIT-managing emotions branch (p = 0.038, 0.025, 0.046, 0.036, 0.025, 0.027, 0.035, 0.028, respectively). G/G homozygotes performed better than T/T in category fluency subtest (p = 0.049). A/A homozygotes of rs3952 performed better than G/G in trail making A subtest (p = 0.048). To our knowledge, this is the first study to indicate that PICK1 polymorphisms may associate with cognitive functions in schizophrenic patients. Further replication studies in healthy controls or other ethnic groups are warranted.