Methyl ß-Cyclodextrin-sperm-mediated gene editing (MBCD-SMGE): a simple and efficient method for targeted mutant mouse production.

BACKGROUND: Generating targeted mutant mice is a crucial technology in biomedical research. This study focuses on optimizing the CRISPR/Cas9 system uptake into sperm cells using the methyl ß-cyclodextrin-sperm-mediated gene transfer (MBCD-SMGT) technique to generate targeted mutant blastocysts and mice efficiently. Additionally, the present study elucidates the roles of cholesterol and reactive oxygen species (ROS) in the exogenous DNA uptake by sperm. RESULTS: In this study, B6D2F1 mouse sperm were incubated in the c-TYH medium with different concentrations of MBCD (0, 0.75, 1, and 2 mM) in the presence of 20 ng/µl pCAG-eCas9-GFP-U6-gRNA (pgRNA-Cas9) for 30 min. Functional parameters, extracellular ROS, and the copy numbers of internalized plasmid per sperm cell were evaluated. Subsequently, in vitro fertilization (IVF) was performed and fertilization rate, early embryonic development, and transfection rate were assessed. Finally, our study investigated the potential of the MBCD-SMGT technique in combination with the CRISPR-Cas9 system, referred to as MBCD-SMGE (MBCD-sperm-mediated gene editing), for generating targeted mutant blastocysts and mice. Results indicated that cholesterol removal from the sperm membrane using MBCD resulted in a premature acrosomal reaction, an increase in extracellular ROS levels, and a dose-dependent influence on the copy numbers of the internalized plasmids per sperm cell. Moreover, the MBCD-SMGT technique led to a larger population of transfected motile sperm and a higher production rate of GFP-positive blastocysts. Additionally, the current study validated the targeted indel in blastocyst and mouse derived from MBCD-SMGE technique. CONCLUSION: Overall, this study highlights the significant potential of the MBCD-SMGE technique for generating targeted mutant mice. It holds enormous promise for modeling human diseases and improving desirable traits in animals.

ZIF-8 Nanoparticle: A Valuable Tool for Improving Gene Delivery in Sperm-Mediated Gene Transfer.

Metal-organic frameworks (MOFs) are porous materials with unique characteristics that make them well-suited for drug delivery and gene therapy applications. Among the MOFs, zeolitic imidazolate framework-8 (ZIF-8) has emerged as a promising candidate for delivering exogenous DNA into cells. However, the potential of ZIF-8 as a vector for sperm-mediated gene transfer (SMGT) has not yet been thoroughly explored.This investigation aimed to explore the potential of ZIF-8 as a vector for enhancing genetic transfer and transgenesis rates by delivering exogenous DNA into sperm cells. To test this hypothesis, we employed ZIF-8 to deliver a plasmid expressing green fluorescent protein (GFP) into mouse sperm cells and evaluated the efficiency of DNA uptake. Our findings demonstrate that ZIF-8 can efficiently load and deliver exogenous DNA into mouse sperm cells, increasing GFP expression in vitro. These results suggest that ZIF-8 is a valuable tool for enhancing genetic transfer in SMGT, with important implications for developing genetically modified animals for research and commercial purposes. Additionally, our study highlights the potential of ZIF-8 as a novel class of vectors for gene delivery in reproductive biology.Overall, our study provides a foundation for further research into using ZIF-8 and other MOFs as gene delivery systems in reproductive biology and underscores the potential of these materials as promising vectors for gene therapy and drug delivery.

Lupus mice derived mesenchymal stromal cells: Beneficial or detrimental on SLE disease outcome.

BACKGROUND: Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disease characterized by the presence of autoantibodies against nuclear genes, deposition of immune complexes, and autoimmune T cells, through which, tissue damage would ultimately occur. Furthermore, loss of immune tolerance and imbalance of Th1/Th2 cells in addition to Th17/Treg are contributed to the pathogenesis of SLE. Mesenchymal stromal cells (MSCs) infusion is a potential therapy for SLE disease. Despite a majority of SLE patients achieving clinical remission after allogeneic MSC infusion from healthy individuals, SLE patients have less benefited from autologous MSC infusion, justifying the probable compromised function of SLE patients-derived MSCs. In this study, we aim to further investigate the potential immunoregulatory mechanisms in which mesenchymal stromal cells derived from pristane-induced lupus mice, following injection into healthy and lupus mice, exert their possible effects on the lupus process. METHOD: 40 female Balb/c mice aged 3 weeks were purchased and randomly divided into six groups. First, lupus disease was induced into the lupus groups by intraperitoneal injection of pristane and then the mice were surveyed for 6 months. The body weight, anti-dsDNA autoantibody levels, serum creatinine, and Blood Urea Nitrogen (BUN) levels were measured in two-month intervals. After 6 months, the group of lupus mice was sacrificed, and lupus MSCs were isolated. Two months later, cultured lupus MSCs were intravenously injected into two groups of healthy and lupus mice. After two months, the mice were euthanized and the kidneys of each group were examined histologically by hematoxylin & eosin (H&E) staining and the immunofluorescence method was also performed to evaluate IgG and C3 deposition. The frequency of splenic Th1, Th2, Th17, and Treg cells was measured by flow cytometry. Moreover, the cytokine levels of IFN-γ, IL-4, IL-17, and TGF-ß in sera were measured by ELISA method. RESULTS: Our results showed that the induction of lupus disease by pristane in Balb/c mice caused the formation of lipogranuloma, increased levels of anti-dsDNA autoantibodies, and impaired renal function in all pristane-induced lupus groups. In addition, the injection of lupus mesenchymal stromal cells (L-MSC) into healthy and lupus mice led to a further rise in anti-dsDNA serum levels, IgG and C3 deposition, and further dysfunction of mice renal tissue. Also, the flow cytometry results implicated that compared to the control groups, splenic Th1, Th2, and Th17 inflammatory cell subtypes and their secreted cytokines (IFN-γ, IL-4, and IL-17) in the sera of healthy and lupus mice were increased after the intake of L-MSC. Additionally, the splenic Treg cells were also significantly increased in the lupus mice receiving L-MSC. However, a decrease in serum levels of TGF-ß cytokine was observed in healthy and lupus mice following L-MSC injection. In contrast, the lupus mice receiving healthy mesenchymal stem cells (H-MSC) manifested opposite results. CONCLUSION: In a nutshell, our results suggest that although allogeneic MSCs are encouraging candidates for SLE treatment, syngeneic MSCs may not be eligible for treating SLE patients due to their defects in regulating the immune system in addition to their capability in promoting inflammation which would consequently worsen the SLE disease status.

Long-term outcomes and prognostic factors of surgical treatment of peri-implantitis - A retrospective study.

AIM: To evaluate long-term outcomes and prognostic factors of non-reconstructive surgical treatment of peri-implantitis. MATERIALS AND METHODS: One hundred forty-nine patients (267 implants) were surgically treated for peri-implantitis and followed for an average of 7.0 (SD: 3.6) years. The primary outcome was implant loss. Additional bone loss and surgical retreatment were secondary outcomes. Patient/implant characteristics, as well as clinical and radiographic parameters collected prior to initial surgery, were evaluated as potential predictors of implant loss. Flexible parametric survival models using restricted cubic spline functions were used; 5- and 10-year predicted rates of implant loss were calculated according to different scenarios. RESULTS: Fifty-three implants (19.9%) in 35 patients (23.5%) were lost during the observation period. Implant loss occurred after a mean period of 4.4 (SD: 3.0) years and was predicted by implant surface characteristics (modified surface; HR 4.5), implant length (HR 0.8 by mm), suppuration at baseline (HR 2.7) and disease severity (baseline bone loss: HR 1.2 by mm). Estimates of 5- and 10-year implant loss ranged from 1% (best prognostic scenario; initial bone loss <40% of implant length, turned implant surface and absence of suppuration on probing (SoP)) to 63% (worst prognostic scenario; initial bone loss ≥60% of implant length, modified implant surface and SoP) and from 3% to 89%, respectively. Surgical retreatment was performed at 65 implants (24.3%) in 36 patients (24.2%) after a mean time period of 4.5 (3.1) years. In all, 59.5% of implants showed additional bone loss, were surgically retreated or lost. CONCLUSIONS: Recurrence of disease is common following surgical treatment of peri-implantitis. The strongest predictor for implant loss was implant surface characteristics. Implant length as well as suppuration and disease severity at baseline were also relevant factors.

A new generation of mesenchymal stromal/stem cells differentially trained by immunoregulatory probiotics in a lupus microenvironment.

BACKGROUND: Increasing evidence suggests that multipotent mesenchymal stem/stromal cells (MSCs) are a promising intervention strategy in treating autoimmune inflammatory diseases. It should be stated that systemic immunoregulation is increasingly recognized among the beneficial effects of MSCs and probiotics in treating morbid autoimmune disorders such as lupus. This study aimed to determine if immunoregulatory probiotics L. rhamnosus or L. delbrueckii can change the immunomodulatory effects of MSCs in lupus-like disease. METHODS: Pristane-induced lupus (PIL) mice model was created via intraperitoneal injection of Pristane and then confirmed. Naïve MSCs (N-MSCs) were coincubated with two Lactobacillus strains, rhamnosus (R-MSCs) or delbrueckii (D-MSCs), and/or a combination of both (DR-MSCs) for 48 h, then administrated intravenously in separate groups. Negative (PBS-treated normal mice) and positive control groups (PBS-treated lupus mice) were also investigated. At the end of the study, flow cytometry and enzyme-linked immunosorbent assay (ELISA) analysis were used to determine the percentage of Th cell subpopulations in splenocytes and the level of their master cytokines in sera, respectively. Moreover, lupus nephritis was investigated and compared. Analysis of variance (ANOVA) was used for multiple comparisons. RESULTS: Abnormalities in serum levels of anti-dsDNA antibodies, creatinine, and urine proteinuria were significantly suppressed by MSCs transplantation, whereas engrafted MSCs coincubation with both L. strains did a lesser effect on anti-dsDNA antibodies. L. rhamnosus significantly escalated the ability of MSCs to scale down the inflammatory cytokines (IFN-É£, IL-17), while L. delbrueckii significantly elevated the capacity of MSCs to scale down the percentage of Th cell subpopulations. However, incubation with both strains induced MSCs with augmented capacity in introducing inflammatory cytokines (IFN-É£, IL-17). Strikingly, R-MSCs directly restored the serum level of TGF-ß more effectively and showed more significant improvement in disease parameters than N-MSCs. These results suggest that R-MSCs significantly attenuate lupus disease by further skew the immune phenotype of MSCs toward increased immunoregulation. CONCLUSIONS: Results demonstrated that Lactobacillus strains showed different capabilities in training/inducing new abilities in MSCs, in such a way that pretreated MSCs with L. rhamnosus might benefit the treatment of lupus-like symptoms, given their desirable properties.

Radon measurement and age-independent effective dose attributed to ingestion of bottled water in Iran: sensitivity analysis.

A comprehensive study was made to measure the radon concentration in bottled water available in Iran market. The 222Rn concentration in 70 bottled water samples were measured by the sniffing mode technique and RTM 1688-2 (SARAD, Germany) in immediate sampling time and 3 months later for determination of radon decay. The measured radon concentration ranged from 0.003 to 0.618 Bq L-1 in bottled water samples, which were much lower than the recommended value for radon in drinking water by WHO (100 Bq L-1) and United states environmental protection agency (USEPA) (11.1 Bq L-1). The annual effective dose of 222Rn due to ingestion bottled water was also evaluated in this research. The mean annual effective dose due to ingestion of radon in bottled water for adults, children, and infants were estimated to vary from 5.30 × 10-4 mSv-1, 4.90 × 10-4 mSv-1, and 2.15 × 10-4 mSv-1, respectively. Overall, this study indicated that the Iranian people receive no significant radiological risk due to exposure to radon concentration in bottled water brands common consumed in Iranian market.

The Value of Strain Echocardiography in Predicting Electrical Progression in Patients With Arrhythmogenic Right Ventricular Cardiomyopathy.

BACKGROUND: Arrhythmogenic right ventricular (RV) cardiomyopathy is a progressive disease characterized by the replacement of the normal myocardium with fibrofatty tissue. This study aimed to determine the value of echocardiographic RV deformation parameters in predicting electrical progression as assessed by serial changes in RV lead sensing and threshold in patients with arrhythmogenic RV cardiomyopathy. METHODS: The present study recruited 40 patients with a definitive diagnosis of arrhythmogenic RV cardiomyopathy at a mean (SD) age of 38.6 (14.2) years between 2018 and 2020. All patients had received an implantable cardioverter-defibrillator for the primary or secondary prevention of sudden cardiac death. The patients underwent 2-dimensional (2D) and 3-dimensional (3D) transthoracic echocardiographic examinations and RV 2D and 3D strain analyses, comprising free-wall longitudinal strain, global longitudinal strain, and strain rate. They were then followed up for electrical progression. RESULTS: During a mean (SD) follow-up period of 20 (6) months, the RV lead amplitude decreased from 7.95 (IQR, 4.53-10.25) mV to 5.25 (IQR, 2.88-8.55) mV (P < .001), and the lead threshold increased from 0.75 (IQR, 0.50-0.79) V to 0.75 (IQR, 0.75-1.00) V (P < .001). Right ventricular 2D free-wall (ρ = 0.56, P = .01), RV 2D global (ρ = 0.58, P = .007), and RV 3D free-wall (ρ = 0.65; P = .003) longitudinal strain correlated with electrical progression. CONCLUSION: Right ventricular 2D and 3D deformation parameters were found to be significant predictors of electrical progression during follow-up of patients with arrhythmogenic RV cardiomyopathy. These findings suggest that echocardiography has a pivotal role in predicting patients at high risk for electrical progression.

Enhanced Ionic Polymer-Metal Composites with Nanocomposite Electrodes for Restoring Eyelid Movement of Patients with Ptosis.

The present study aims to use enhanced ionic polymer-metal composites (IPMC) as an artificial muscle (a soft-active actuator) to restore eyelid movement of patients with ptosis. The previous eyelid movement mechanisms contained drawbacks, specifically in the lower eyelid. We used finite element analysis (FEA) to find the optimal mechanism among two different models (A and B). In addition to common electrodes of IPMC (gold and platinum), the bovine serum albumin (BSA) and microcrystalline cellulose (MCC) polymers, with optimal weight percentages of carbon nanotube (CNT) nanofiller, were also utilized as non-metallic electrodes to improve the efficiency of the IPMC actuator. In both models, IPMC with nanocomposite electrodes had higher efficiency as compared to the metallic electrodes. In model A, which moved eyelids indirectly, IPMC with MCC-CNT electrode generated a higher force (25.4%) and less stress (5.9 times) as compared to IPMC with BSA-CNT electrode. However, the use of model A (even with IPMCs) with nanocomposite electrodes can have limitations such as possible malposition issues in the eyelids (especially lower). IPMC with MCC-CNT nanocomposite electrode under model B, which moved eyelids directly, was the most efficient option to restore eyelid movement. It led to higher displacements and lower mechanical stress damage as compared to the BSA-CNT. This finding may provide surgeons with valuable data to open a window in the treatment of patients with ptosis.

Effects of human chorionic gonadotropin-producing peripheral blood mononuclear cells on the endometrial receptivity and implantation sites of the mouse uterus.

OBJECTIVE: This research investigated the effects of human chorionic gonadotropin (HCG)-producing peripheral blood mononuclear cells (PBMCs) on the implantation rate and embryo attachment in mice. METHODS: In this experimental study, a DNA fragment of the HCG gene was cloned into an expression vector, which was transfected into PBMCs. The concentration of the produced HCG was measured using enzyme-linked immunosorbent assay. Embryo attachment was investigated on the co-cultured endometrial cells and PBMCs in vitro. As an in vivo experiment, intrauterine administration of PBMCs was done in plaque-positive female mice. Studied mice were distributed into five groups: control, embryo implantation dysfunction (EID), EID with produced HCG, EID with PBMCs, and EID with HCG-producing PBMCs. Uterine horns were excised to characterize the number of implantation sites and pregnancy rate on day 7.5 post-coitum. During an implantation window, the mRNA expression of genes was evaluated using real-time polymerase chain reaction. RESULTS: DNA fragments were cloned between the BamHI and EcoRI sites in the vector. About 465 pg/mL of HCG was produced in the transfected PBMCs. The attachment rate, pregnancy rate, and the number of implantation sites were substantially higher in the HCG-producing PBMCs group than in the other groups. Significantly elevated expression of the target genes was observed in the EID with HCG-producing PBMCs group. CONCLUSION: Alterations in gene expression following the intrauterine injection of HCG-producing PBMCs, could be considered a possible cause of increased embryo attachment rate, pregnancy rate, and the number of implantation sites.

Production of CFTR Mutant Gene Model by Homologous Recombination System.

OBJECTIVE: The most common mutation in cystic fibrosis (CF), (ΔF508-CFTR), results in impaired protein maturation, folding and transportation to the surface of the cell. As a consequence of impaired protein maturation and/or transport from the extracellular matrix to the cell, different systems are influenced, including gastrointestinal system and glandular system, reproductive system and respiratory systems. CF models are essential tools to provide further knowledge of CF pathophysiology. With this aim, we designed a transgenic CF model based on the homologous recombination (HR) system. MATERIALS AND METHODS: In this experimental study, a specifically designed construct containing the CFTR gene with F508del was cloned into a PTZ57R cloning vector and then the construct was transformed into the male pronucleus by microinjection after in vitro fertilization (IVF). Then the rates of blastocyst formation and embryonic development at 72 hours after IVF, were evaluated using the inverted microscope and the insertion of the construct was approved by polymerase chain reaction (PCR) method. RESULTS: The CFTR gene was successfully cloned into the PTZ57R cloning vector and overall, from 22 injected cells, 5 blastocysts were observed after pronuclear injection of the CFTR gene construct. PCR verification of the blastocyst with CFTR-specific primers represented complete recombination of CFTR into the mouse genome. CONCLUSION: For the first time we designed a unique genome construction that can be detected using a simple PCR method. The pronuclear injection was performed for the transformation of the genome construct into the male pronuclei using microinjection and the development of zygote to the blastocyst stage has been observed following transgenesis.

Tissue Doppler echocardiography and outcome in arrhythmogenic right ventricular cardiomyopathy.

AIMS: This study aimed at investigating whether tissue Doppler imaging (TDI) is associated with adverse events in arrhythmogenic right ventricular cardiomyopathy (ARVC). METHODS AND RESULTS: Transthoracic echocardiography was performed in 72 patients with definite (n = 63) or borderline (n = 9) ARVC diagnosed according to the 2010 Task Force Criteria and included in the prospective Zurich ARVC registry. Myocardial peak systolic tissue velocity (S') was measured by TDI at lateral tricuspid (tricuspid S'), medial mitral (septal S'), and lateral mitral annulus (lateral S'). Association of echocardiographic parameters with outcome was assessed by univariable Cox regression. During a median follow-up of 4.9 ± 2.6 years, 6 (8.3%) patients died of cardiovascular cause or received heart transplantation and 21 (29.2%) patients developed sustained ventricular arrhythmia. Tricuspid, septal, and lateral S' were lower in patients who died (p = 0.001; p < 0.001; p = 0.008; respectively), while tricuspid and septal S' were lower in those with ventricular arrhythmia (p = 0.001; p = 0.008; respectively). There was a significant association of tricuspid, septal, and lateral S' with mortality (HR = 1.61, p = 0.011; HR = 2.15, p = 0.007; HR = 1.67, p = 0.017; respectively), while tricuspid and septal S' were associated with ventricular arrhythmia (HR = 1.20, p = 0.022; HR = 1.37, p = 0.004; respectively). Kaplan-Meier analyses demonstrated a higher freedom from mortality with tricuspid S' >8 cm/s (p = 0.001) and from ventricular arrhythmia with S' >10.5 cm/s (p = 0.021). CONCLUSIONS: This study demonstrates that TDI provides information on the ARVC phenotype, is associated with adverse events in ARVC patients, and differentiates between patients with and without adverse events.

Fluoride removal from aqueous solution via environmentally friendly adsorbent derived from seashell.

Nowadays, the presence of excessive ions in water resources is of utmost concern and has attracted increasing attention; therefore, excessive amounts of these ions such as fluoride should be removed from drinking water. Conventional water treatment processes are shown to be incapable of the complete removal of redundant fluoride from aqueous water bodies, whereas adsorption is a promising, effective, cost-benefit, and simple method for this purpose. This study aimed to synthesize effective adsorbents from bivalve shells and evaluate the adsorption function of bivalve shells in removing fluoride from aqueous solutions. In this study, the oyster shell was collected from the Persian Gulf's seaside and were crushed by manual mortar and blender, and graded with standard sieves with 70 mesh size. The prepared bivalve shell was characterized by SEM and FTIR. To investigate and optimize various variables on fluoride removal percentage a response surface methodology based on central composite design (RSM-CCD) was used. Under optimal conditions (pH: 5.5, adsorbent dose: 0.3 g/L, contact time: 85 min and fluoride concentration: 3 mg/L) the maximum removal efficiency was 97.26%. Results showed that the adsorption equilibrium and kinetic data were matched with the isotherm Langmuir Model (R2 = 0.98) with qmax = 27.31 mg/g and pseudo-second-order reaction (R2 = 0.99). Also, a thermodynamic study exhibited that the adsorption process of fluoride into bivalve shells was an exothermic reaction and could not be a spontaneous adsorption process. Based on the results, the bivalve shell was found as an appropriate adsorbent to remove fluoride from aqueous solutions.

Right atrial strain and cardiovascular outcome in arrhythmogenic right ventricular cardiomyopathy.

AIMS: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is characterized by progressive fibro-fatty infiltration of the myocardium and associated with adverse cardiovascular (CV) events. This study aims to examine right atrial (RA) deformation in ARVC and understand its association with CV outcomes. METHODS AND RESULTS: RA strain was determined in 50 patients with definite ARVC, compared with a matched control group of 50 healthy individuals, and analysed for outcome association over a median follow-up duration of 5 years. A subgroup of 30 ARVC patients with normal RA volume (ARVC-N group) was compared with 30 matched controls (Control-N), and the outcome was analysed separately. RA reservoir, conduit, and pump strain were significantly impaired in ARVC vs. control. Similar observations were made in the N-ARVC subgroup. Reservoir strain was associated with an increased risk of atrial arrhythmia (AA) [hazard ratio (HR) 0.88, P < 0.01] and CV events (HR 0.92, P < 0.01). Conduit strain also predicted AA (HR 1.02, P < 0.01), while pump strain predicted CV events (HR 1.09, P = 0.02). Reservoir strain improved the fitness of bivariable models for the association of RV end-diastolic area index, RV fractional area change, and RV global longitudinal strain with CV events. CONCLUSION: ARVC patients display impaired RA strain even when RA volume is normal. Reservoir and pump strain are associated with an increased risk of CV events. Reservoir strain improved model fitness for the association of RVGLS and other echocardiographic parameters with CV events.

The Use of Trichostatin A during Pluripotent Stem Cell Generation Does Not Affect MHC Expression Level.

Pluripotent stem cells (PSCs) are considered as a potent tool for use in regenerative medicine. Highly efficient generation of PSCs through chromatin modulators such as trichostatin A (TSA) might change their MHC molecule expression profile. The efficiency of PSC generation and their immunogenicity is major obstacles for clinical use. Hence, we aim to investigate whether the use of TSA during PSC generation affects MHC expression level. Three PSC lines were generated by iPSCs, NT-ESCs, and IVF-ESCs' reprogramming methods from B6D2F1 mouse embryonic fibroblast cells. Established PSC lines were characterized by alkaline phosphatase assay (ALP) and immunocytochemistry. Their chromosome fidelity was checked by karyotyping. The expression level of pluripotent genes (oct4, nanog, sox2, klf4), HDACs (hdac1, hdac2, and hdac3), and immune-related genes (including Qa-1, Qa-2, H2kb, H2kd, H2db, H2db, CIITA, H2-IE-ßb, H2-IE-ßd) in iPSC and ESC lines were assessed by real-time PCR analysis. The presence of MHC molecules on the surface of pluripotent stem cells was also checked by flow cytometry technique. Significant increase of pluripotency markers, oct4, nanog, sox2, and klf4, was observed in 100 nM TSA-treated samples. 100 nM TSA induced significant upregulation of H2db in generated iPSCs. H2-IE-ßd was remarkably downregulated in 50 and 100 nM TSA-treated iPSC lines. The expression level of other immune-related genes was not greatly affected by TSA in iPSC and NT-ESC lines. It is concluded that the use of short-term and low concentration of TSA during reprogramming in PSC generation procedure significantly increases PSC generation efficiency, but do not affect the MHC expression in established cell lines, which is in the benefit of cell transplantation in regenerative medicine.

Left ventricular twist predicts mortality in severe aortic stenosis.

OBJECTIVE: Left ventricular (LV) twist is a major component of ventricular mechanics reflecting the helical orientation of cardiac fibres and compensating for afterload mismatch. However, it is not known whether it determines outcome after transcatheter aortic valve implantation (TAVI). This study sought to investigate TAVI-induced short-term changes of LV twist and to define its role in outcome prediction. METHODS: A total of 146 patients (median age 81.78 years, 50.7% male) undergoing TAVI for severe aortic stenosis were included. LV rotation and twist were determined by speckle tracking echocardiography within 3 months before and 2 weeks after TAVI. All-cause mortality at 2 years was defined as primary end point. RESULTS: Patients who survived exhibited a higher apical peak systolic rotation (APSR) (p<0.001), twist (p=0.003) and torsion (p=0.019) pre-TAVI compared with those who died (n=22). Within 2 weeks after TAVI, APSR, twist and torsion decreased in patients who survived (all p<0.001), while no change occurred in those who died. Cox regression analysis showed an association of pre-TAVI APSR (HR 0.92, p=0.010), twist (HR 0.93, p=0.018) and torsion (HR 0.68, p=0.040) with all-cause mortality and an even stronger association of the respective changes after TAVI (∆APSR: HR 1.15, p<0.001; ∆twist: HR 1.14, p<0.001; ∆torsion: HR 2.53, p<0.001). All the parameters determined outcome independently of global longitudinal strain (GLS) and LV ejection fraction (LVEF). CONCLUSION: APSR, twist and torsion pre-TAVI as well as their change within 2 weeks after TAVI predict 2-year all-cause mortality after TAVI, adding incremental prognostic value to LVEF and GLS.

Tricostatin A-treated round spermatid enhances preimplantation embryo developmental competency following round spermatid injection in mice.

It has been documented that the inefficacy of round spermatid injection (ROSI) might be caused by abnormal epigenetic modifications. Therefore, this study aimed to evaluate the effect of trichostatin A (TSA) as an epigenetic modifier of preimplantation embryo development in activated ROSI oocytes. Matured oocytes were collected from superovulated female mice. Testes were placed in human tubal fluid medium and masses were then cut into small pieces to disperse spermatogenic cells. Round spermatids were treated with TSA and subsequently injected into oocytes. The expression level of the development-related genes including Oct4, Sox2, Nanog, Dnmt and Hdac transcripts were evaluated using qRT-PCR. Immunohistochemistry was performed to confirm the presence of Oct-4 protein at the blastocyst stage. There was no statistically significant difference in fertilization rate following ROSI/+TSA compared with the non-treated ROSI and intracytoplasmic sperm injection (ICSI) groups. Importantly, TSA treatment increased blastocyst formation from 38% in non-treated ROSI to 68%. The relative expression level of developmentally related genes increased and Dnmt transcripts decreased in ROSI/+TSA-derived embryos, similar to the expression levels observed in the ICSI-derived embryos. In conclusion, our results indicate that spermatid treatment with TSA prior to ROSI would increase the success rate of development to the blastocyst stage and proportion of pluripotent cells.

Predictors of worsening TR severity after right ventricular lead placement: any added value by post-procedural fluoroscopy versus three -dimensional echocardiography?

BACKGROUND: The effect of right ventricular (RV) leads on tricuspid valve has been already raised concerns, especially in terms of prognostic implication. For such assessment, three-dimensional transthoracic echocardiography (3D-TTE) has been used previously but there was no data on the use of post-procedural fluoroscopy in the literature. METHODS: We prospectively enrolled 59 patients who underwent clinically indicated placement of pacemaker or implantable cardioverter defibrillator (ICD). Vena contracta (VC) and tricuspid regurgitation (TR) severity were measured using two-dimensional transthoracic echocardiography (2D-TTE) at baseline. Follow up 3D-TTE was performed 6 months after device implantation to assess TR severity and RV lead location. RESULTS: Lead placement position in TV was defined in 51 cases.TR VC was increased after the lead placement, compared to the baseline study (VC: 3.86 ± 2.32 vs 3.18 ± 2.39; p = 0.005), with one grade worsening in TR in 25.4% of cases. The mean changes in VC levels were 1.14 ± 0.67 mm. Among all investigated parameters, VC changes were predicted based on lead placement position only in 3D-TTE (p < 0.001) while the other variables including fluoroscopy parameters were not informative. CONCLUSION: The RV Lead location examined by 3D-TTE seems to be a valuable parameter to predict the changes in the severity of the tricuspid regurgitation. Fluoroscopy findings did not improve the predictive performance, at least in short term follow up.

Meiosis Resumption of Immature Human Oocytes following Treatment with Calcium Ionophore In Vitro.

OBJECTIVE: In vitro maturation (IVM) of human oocytes is used to induce meiosis progression in immature retrieved oocytes. Calcium (Ca2+) has a central role in oocyte physiology. Passage through meiosis phase to another phase is controlled by increasing intracellular Ca2+. Therefore, the current research was conducted to evaluate the role of calcium ionophore (CI) on human oocyte IVM. MATERIALS AND METHODS: In this clinical trial study, immature human oocytes were obtained from 216 intracytoplasmic sperm injection (ICSI) cycles. After ovarian stimulation, germinal vesicle (GV) stage oocytes were collected and categorized into two groups: with and without 10 µM CI treatment. Next, oocyte nuclear maturation was assessed after 24-28 hours of culture. Real-time reverse transcription polymerase chain reaction (RT-PCR) was used to assess the transcript profile of several oocyte maturation-related genes (MAPK3, CCNB1, CDK1, and cyclin D1 [CCND1]) and apoptotic-related genes (BCL-2, BAX, and Caspase-3). Oocyte glutathione (GSH) and reactive oxygen species (ROS) levels were assessed using Cell Tracker Blue and 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) fluorescent dye staining. Oocyte spindle configuration and chromosome alignment were analysed by immunocytochemistry. RESULTS: The metaphase II (MII) oocyte rate was higher in CI-treated oocytes (73.53%) compared to the control (67.43%) group, but this difference was not statistically significant (P=0.13). The mRNA expression profile of oocyte maturation-related genes (MAPK3, CCNB1, CDK1, and CCND1) (P<0.05) and the anti-apoptotic BCL-2 gene was remarkably up-regulated after treatment with CI (P=0.001). The pro-apoptotic BAX and Caspase-3 relative expression levels did not change significantly. The CI-treated oocyte cytoplasm had significantly higher GSH and lower ROS (P<0.05). There was no statistically significant difference in meiotic spindle assembly and chromosome alignment between CI treatment and the control group oocytes. CONCLUSION: The finding of the current study supports the role of CI in meiosis resumption of human oocytes. (Registration Number: IRCT20140707018381N4).

Syringic Acid Attenuates Cardiomyopathy in Streptozotocin-Induced Diabetic Rats.

OBJECTIVES: Diabetic cardiomyopathy (DC) has become one of the serious complications in diabetic cases. In this study, we aimed to explore the syringic acid (SYR) protective effect against diabetes-induced cardiac injury in experimental rats. METHODS: Rats were divided in control and streptozotocin-induced diabetic rats which were subdivided into diabetic controls, and three test groups (SYR at 25, 50, and 100 mg/kg) and the nondiabetic group received 100 mg/kg of SYR. All treatments were given SYR for 6 weeks. SYR effects on cardiac diagnostic markers, heart lipid peroxidation, protein carbonylation, antioxidant system, and changes of the heart mitochondrial mass and biogenesis were measured. RESULTS: Diabetes induction prompted CK-MB, LDH levels in serum, cardiac catalase, and superoxide dismutase activity, as well as cardiac TBARs and carbonylated protein. SYR administration (100 m/kg) attenuated CK-MB and LDH levels. Also, 50 and 100 mg/kg of SYR reduced cardiac TBARs and carbonylated protein in diabetic rats. These treatments did not show any effects on GSH content, mtDNA, and mitochondrial biogenesis indices (PGC1- α, NRF1, NRF2, and TFAM) in heart tissue. CONCLUSIONS: SYR treatment showed protective effects on diabetic cardiomyopathy in rats by reducing lipid peroxidation and protein carbonylation. The possible mechanisms could be related to antioxidant activity of this phenolic acid. SYR might play a role of a protective factor in cardiac challenges in diabetes.