Immunosuppression with cyclosporine versus tacrolimus shows distinctive nephrotoxicity profiles within renal compartments.

AIM: Calcineurin inhibitors (CNIs) are the backbone for immunosuppression after solid organ transplantation. Although successful in preventing kidney transplant rejection, their nephrotoxic side effects contribute to allograft injury. Renal parenchymal lesions occur for cyclosporine A (CsA) as well as for the currently favored tacrolimus (Tac). We aimed to study whether chronic CsA and Tac exposures, before reaching irreversible nephrotoxic damage, affect renal compartments differentially and whether related pathogenic mechanisms can be identified. METHODS: CsA and Tac were administered chronically in wild type Wistar rats using osmotic minipumps over 4 weeks. Functional parameters were controlled. Electron microscopy, confocal, and 3D-structured illumination microscopy were used for histopathology. Clinical translatability was tested in human renal biopsies. Standard biochemical, RNA-seq, and proteomic technologies were applied to identify implicated molecular pathways. RESULTS: Both drugs caused significant albeit differential damage in vasculature and nephron. The glomerular filtration barrier was more affected by Tac than by CsA, showing prominent deteriorations in endothelium and podocytes along with impaired VEGF/VEGFR2 signaling and podocyte-specific gene expression. By contrast, proximal tubule epithelia were more severely affected by CsA than by Tac, revealing lysosomal dysfunction, enhanced apoptosis, impaired proteostasis and oxidative stress. Lesion characteristics were confirmed in human renal biopsies. CONCLUSION: We conclude that pathogenetic alterations in the renal compartments are specific for either treatment. Considering translation to the clinical setting, CNI choice should reflect individual risk factors for renal vasculature and tubular epithelia. As a step in this direction, we share protein signatures identified from multiomics with potential pathognomonic relevance.

Propolis-Loaded Poly(lactic-co-glycolic Acid) Nanofibers: An In Vitro Study.

Nanofibers have high potential through their high porosity, small pore sizes, lightweight materials, and their ability to mimic the extracellular matrix structure for use in the manufacture of wound dressings for wound treatment. In this study, poly(lactic-co-glycolic acid) (PLGA) nanofibers were produced by electrospinning. Propolis was loaded into the PLGA nanofibers by the dropping method. The average diameters and effects of propolis loading on the morphology of 37.5, 50, and 100% propolis-loaded PLGA nanofibers (PLGA-P37.5, PLGA-P50, and PLGA-P100) were evaluated by scanning electron microscopy (SEM). The successful loading of propolis into PLGA nanofibers was confirmed with Fourier transform infrared spectroscopy (FTIR) analysis. In vitro propolis release was examined at physiological pH. The antioxidant activity of propolis-loaded nanofibers was studied with 2,2-diphenyl-1-picrylhydrazyl (DPPH). Antimicrobial activities of the nanofibers against Escherichia coli, Staphylococcus aureus and Candida albicans strains were determined by the disk diffusion method. Consequently, PLGA-P50 and PLGA-P100 showed high antimicrobial activity on S. aureus and C. albicans. Cell viability was tested by 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, and propolis-loaded PLGA nanofibers were found to be biocompatible with human fibroblast cells. In the wound scratch assay, propolis-loaded nanofibers supported wound closure with cell migration and proliferation. Thus, in vitro wound closure properties of propolis-loaded PLGA nanofibers were evaluated for the first time in the literature.

Furosemide rescues hypercalciuria in familial hypomagnesaemia with hypercalciuria and nephrocalcinosis model.

AIM: Perturbed calcium homeostasis limits life expectancy in familial hypomagnesaemia with hypercalciuria and nephrocalcinosis (FHHNC). This rare disease occurs by loss-of-function mutations in CLDN16 or CLDN19 genes, causing impaired paracellular reabsorption of divalent cations along the cortical thick ascending limb (cTAL). Only partial compensation takes place in the ensuing late distal convoluted tubule, connecting tubule, and collecting duct, where the luminal transient receptor potential channel V5 (TRPV5), as well as basolateral plasma membrane calcium ATPase (PMCA) and sodium-potassium exchanger (NCX1) mediate transcellular Ca2+ reabsorption. The loop diuretic furosemide induces compensatory activation in these distal segments. Normally, furosemide enhances urinary calcium excretion via inhibition of the aforementioned cTAL. As Ca2+ reabsorption in the cTAL is already severely impaired in FHHNC patients, furosemide may alleviate hypercalciuria in this disease by activation of the distal transcellular Ca2+ transport proteins. METHODS: Cldn16-deficient mice (Cldn16-/- ) served as a FHHNC model. Wild-type (WT) and Cldn16-/- mice were treated with furosemide (7 days of 40 mg/kg bw) or vehicle. We assessed renal electrolyte handling (metabolic cages) and key divalent transport proteins. RESULTS: Cldn16-/- mice show higher Ca2+ excretion than WT and compensatory stimulation of Cldn2, TRPV5, and NCX1 at baseline. Furosemide reduced hypercalciuria in Cldn16-/- mice and enhanced TRPV5 and PMCA levels in Cldn16-/- but not in WT mice. CONCLUSIONS: Furosemide significantly reduces hypercalciuria, likely via upregulation of luminal and basolateral Ca2+ transport systems in the distal nephron and collecting duct in this model for FHHNC.

XBP1 Activation Reduces Severity of Polycystic Kidney Disease due to a Nontruncating Polycystin-1 Mutation in Mice.

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in Pkd1 and Pkd2. They encode the polytopic integral membrane proteins polycystin-1 (PC1) and polycystin-2 (PC2), respectively, which are expressed on primary cilia. Formation of kidney cysts in ADPKD starts when a somatic second hit mechanism inactivates the wild-type Pkd allele. Approximately one quarter of families with ADPDK due to Pkd1 have germline nonsynonymous amino acid substitution (missense) mutations. A subset of these mutations is hypomorphic, retaining some residual PC1 function. Previous studies have shown that the highly conserved Ire1 α -XBP1 pathway of the unfolded protein response can modulate levels of functional PC1 in the presence of mutations in genes required for post-translational maturation of integral membrane proteins. We examine how activity of the endoplasmic reticulum chaperone-inducing transcription factor XBP1 affects ADPKD in a murine model with missense Pkd1 . METHODS: We engineered a Pkd1 REJ domain missense murine model, Pkd1 R2216W , on the basis of the orthologous human hypomorphic allele Pkd1 R2220W , and examined the effects of transgenic activation of XBP1 on ADPKD progression. RESULTS: Expression of active XBP1 in cultured cells bearing PC1 R2216W mutations increased levels and ciliary trafficking of PC1 R2216W . Mice homozygous for Pkd1 R2216W or heterozygous for Pkd1 R2216Win trans with a conditional Pkd1 fl allele exhibit severe ADPKD following inactivation in neonates or adults. Transgenic expression of spliced XBP1 in tubule segments destined to form cysts reduced cell proliferation and improved Pkd progression, according to structural and functional parameters. CONCLUSIONS: Modulating ER chaperone function through XBP1 activity improved Pkd in a murine model of PC1, suggesting therapeutic targeting of hypomorphic mutations.

Immunosuppressive calcineurin inhibitor cyclosporine A induces proapoptotic endoplasmic reticulum stress in renal tubular cells.

Current immunosuppressive strategies in organ transplantation rely on calcineurin inhibitors cyclosporine A (CsA) or tacrolimus (Tac). Both drugs are nephrotoxic, but CsA has been associated with greater renal damage than Tac. CsA inhibits calcineurin by forming complexes with cyclophilins, whose chaperone function is essential for proteostasis. We hypothesized that stronger toxicity of CsA may be related to suppression of cyclophilins with ensuing endoplasmic reticulum (ER) stress and unfolded protein response (UPR) in kidney epithelia. Effects of CsA and Tac (10 µM for 6 h each) were compared in cultured human embryonic kidney 293 (HEK 293) cells, primary human renal proximal tubule (PT) cells, freshly isolated rat PTs, and knockout HEK 293 cell lines lacking the critical ER stress sensors, protein kinase RNA-like ER kinase or activating transcription factor 6 (ATF6). UPR was evaluated by detection of its key components. Compared with Tac treatment, CsA induced significantly stronger UPR in native cultured cells and isolated PTs. Evaluation of proapoptotic and antiapoptotic markers suggested an enhanced apoptotic rate in CsA-treated cells compared with Tac-treated cells as well. Similar to CsA treatment, knockdown of cyclophilin A or B by siRNA caused proapoptotic UPR, whereas application of the chemical chaperones tauroursodeoxycholic acid or 4-phenylbutyric acid alleviated CsA-induced UPR. Deletion of protein kinase RNA-like ER kinase or ATF6 blunted CsA-induced UPR as well. In summary, inhibition of cyclophilin chaperone function with ensuing ER stress and proapoptotic UPR aggravates CsA toxicity, whereas pharmacological modulation of UPR bears potential to alleviate renal side effects of CsA.

Caffeic Acid Phenethyl Ester Loaded Electrospun Nanofibers for Wound Dressing Application.

Electrospinning is an advantageous method with a wide usage area, which enables the production of materials consisting of nano-thickness fibers. In this study, caffeic acid phenethyl ester (CAPE) molecule was loaded onto the poly(lactic-co-glycolic acid) (PLGA) nanofibers and obtained nanofibers were physicochemically and biologically investigated for the first time in the literature. The existence of CAPE molecules, loaded on PLGA membranes by dropping and spraying methods, was evaluated by a comparative investigation of Fourier-transform infrared (FTIR) spectra and X-Ray diffraction (XRD) patterns. Fiber morphology of the membranes was investigated by scanning electron microscope (SEM). CAPE release and swelling behaviors of the membranes were studied in vitro. The radical scavenging activity of CAPE-loaded wound dressing materials was determined by using an antioxidant assay. The antimicrobial properties of PLGA and CAPE-loaded PLGA membranes were evaluated against S. aureus, P. aeruginosa and C. albicans strains by the time-kill method. The biocompatibility study of the obtained CAPE-loaded fibers conducted on human fibroblast cell line and wound healing promoting effect of the fibers was investigated in vitro scratch assay. The results show that CAPE-loaded PLGA membranes are highly antimicrobial against all strains used in the experiment. Additionally, the results show that they are biocompatible and have wound healing properties on human fibroblasts.

Connexin43 is differentially distributed within renal vasculature and mediates profibrotic differentiation in medullary fibroblasts.

Connexins (Cxs) form gap junctions for intercellular exchange of inorganic ions and messenger molecules. In the kidney, Cxs play essential roles within its compartments, but data on the precise cellular localization and cell type-related function of their isoforms are scarce. We tested whether Cx43 distribution is restricted to vascular and interstitial cells and whether medullary fibroblasts express Cx43 to coordinate profibrotic signaling. Confocal immunofluorescence techniques, ultrastructural labeling, and functional experiments in cell culture were performed. Cx43 was chiefly expressed in the vasculature but was absent from tubular epithelia. All arterial, arteriolar, and lymphatic endothelia showed continuous Cx43 signal along their borders. In the inner medulla, only the interstitium showed Cx43 signals, which were assigned to fibroblasts and their processes. Cultured Cx43-expressing medullary fibroblasts served to study the role of gap junctions in a profibrotic context. In a dye spreading assay, Cx43-sensitive diffusion of Lucifer yellow was dependent on gap junctional passage. The addition of transforming growth factor-ß1 (5 ng/mL for 48 h) activated Cx43 biosynthesis and caused Cx43-sensitive transformation of the fibroblasts into a myofibroblast phenotype. This suggested that Cx43 gap junctional channels enable the coordination of profibrotic signaling between cells of the medullary interstitium. In summary, we demonstrate the presence of Cx43-expressing gap junctions within the two major renal compartments, the vasculature and interstitium. Endothelial Cx43 likely provides functions of an earlier-defined "electrical syncytium" within the vascular wall. Additionally, Cx43 facilitates profibrotic signaling between medullary interstitial fibroblasts.

An Important Tool in Lymphedema Management: Validation of Turkish Version of the Patient Benefit Index-Lymphedema.

Background: The aim of this study was to investigate the Turkish adaptation, validity, and reliability of the Patient Benefit Index-Lymphedema (PBI-L) ensuring direct assessment of the benefit in patients with lymphedema (LE), lipedema, and lipolymphedema. Methods and Results: Eighty-one patients who were consulted for physiotherapy, whose diagnoses were LE, lipedema, and lipolymphedema, and who were treated or planned to be treated for these diagnoses, were included in this study. PBI-L was adapted to Turkish by considering the stages of the cultural adaptation process. Short Form-36 (SF-36) was applied for the validity of PBI-L. PBI-L was repeated after a 1-week interval for test-retest reliability. The mean age was 47.66 ± 14.23 years. The intraclass correlation coefficient (ICC) value was determined as 0.73 (p < 0.0001) for the total score. There was a moderate correlation between first (ICC = 0.63, p < 0.0001) and second subdimensions (ICC = 0.62, p < 0.0001). Cronbach's α values ranged between 0.83 and 0.89. Low correlations were found between total scores of PBI-L and mental health, physical function subdimensions of SF-36 (p < 0.05). The Kaiser Meyer Olkin value was 0.6, and it was found that the PBI-L was not consistent with factor analysis. Conclusion: The Turkish version of PBI-L is a valid and reliable tool in patients with LE, lipedema, and lipolymphedema. However, the reassessment validity of PBI-L would be suggested by using an LE-specific quality of life questionnaire.

Translation and Validation of the Turkish Version of Lymphedema Quality of Life Tool (LYMQOL) in Patients with Breast Cancer Related Lymphedema.

OBJECTIVE: Breast cancer related lymphedema (BCRL) is a drastic situation that affects patients who have undergone breast cancer surgery. The impact of this condition on individuals' quality of life should be investigated in more detail to obtain better treatment results. MATERIALS AND METHODS: In total, 65 patients with BCRL participated in this study. Nottingham Health Profile (NHP) was used to evaluate the validity of associated domains in Lymphedema Quality of Life Tool (LYMQoL). Both the LYMQoL and NHP were filled out by BCRL patients. To evaluate its test-retest reliability, the LYMQoL was subsequently performed seven days following its initial application. Measurement properties such as internal consistency, test-retest reliability, criterion validity and factor structure were tested. The internal consistency was assessed via Cronbach's alpha; test-retest reliability was assessed by the intra-class correlation coefficient (ICC). RESULTS: Cronbach's alpha values ranged from 0.74 to 0.91 for the LYMQoL total and domain scores. Test-retest reliability was excellent (ICC=0.92-0.99). When the relation between LYMQoL and NHP was investigated, 'good' to 'very good' correlations were obtained (r=0.539-0.643, p<0.05) for all domains of LYMQoL. Exploratory factor analyses demonstrated a four-factor structure. CONCLUSION: Turkish version of LYMQoL is a valid and reliable measurement tool to evaluate the quality of life in patients with BCRL.

The effect of pelvic organ prolapse type on sexual function, muscle strength, and pelvic floor symptoms in women: A retrospective study.

OBJECTIVE: This retrospective research was planned to investigate the effect of pelvic organ prolapse (POP) type on sexual function, muscle strength, and pelvic floor symptoms in symptomatic women. MATERIALS AND METHODS: Data on POP type and stages as assessed using the Pelvic Organ Prolapse-Quantification system of 721 women who presented to the women's health unit between 2009 and 2016 were collected retrospectively. POP types were recorded as asymptomatic, anterior, apical, and posterior compartment prolapses. Sexual function was assessed using the Pelvic Organ Prolapse/Urinary Incontinence Sexual Questionnaire short-form (PISQ-12), pelvic floor muscle strength was assessed through vaginal pressure measurement, and pelvic floor symptoms and quality of life were assessed using the Pelvic Floor Distress Inventory-20 (PFDI-20). RESULTS: Among 168 women who met the inclusion criteria, 96 had anterior compartment prolapses, 20 had apical compartment prolapses, 16 had posterior compartment prolapses, and 36 women were asymptomatic. There was no difference between the groups in their PISQ-12 total and subscales scores, PFDI-20 total and two subscale (colorectal/anal, urinary) scores, and muscle strength (p>0.05). In the Pelvic Organ Prolapse Distress Inventory-6, another subscale of PFDI-20, it was determined that there was a difference between asymptomatic women and those with anterior compartment prolapses (p=0.044) and apical compartment prolapses (p=0.011). CONCLUSION: This research found that POP type did not affect sexual function, muscle strength, and colorectal and urinary symptoms in our cohort. There were more prolapse symptoms and complaints in women with anterior and apical compartment prolapses.

The effects of short-term use of compression stockings on health related quality of life in patients with chronic venous insufficiency.

[Purpose] This study was aimed to analyse the effects of short-term use of compression stockings (CS) on symptoms and QoL in patients with Chronic Venous Insufficiency (CVI). [Subjects and Methods] Based on the CEAP classification C2 and C3, 117 patients with CVI were enrolled in this study. The participants were divided into two groups. The control group refused to use CS, however, was advised to do exercises and take skin care whereas the CS group used CS and performed exercise. The data were collected by using Nottingham Health Profile (NHP), Venous Insufficiency Epidemiological and Economic Study (VEINES-QoL/Sym) and Beck Depression Inventory (BDI) at baseline and after four weeks of treatment and compared within and in between groups. [Results] The comparison of pre- and post-treatment differences between groups was statistically significant for all parameters. In the study group, pre- and post-treatment scores for each parameter were significantly different. However, elevated scores in the control group suggested worsening of the illness. [Conclusion] This study established that short-term use of CS in patients with CVI significantly improved disease specific and general QoL by reducing venous symptoms. Further studies with larger sample size are necessary to confirm these findings.