Cryopreservation of blastomeres separated from two-cell mouse embryos by an ultrarapid freezing method

Purpose To clarify the developmental capacity of frozen two-cell blastomeres, we investigated in vivo and in vitro viabilities of blastomeres that were frozen ultrarapidly after separation from two-cell mouse embryos. Two-cell embryos obtained from superovulated F₁ hybrid females were denuded by treatment with 0.5% pronase solution and then induced to separate into two single blastomeres by gentle pipetting. The blastomeres were cryopreserved by an ultrarapid freezing method.Results The preimplantation developmental rate of two-cell embryos frozen in 3.0 MDMSO was significantly higher than the rate of those frozen in 15 and 4.5 MDMSO (at least P<0.05). The in vitro developmental rate of the ultrarapidly frozen-thawed blastomeres separated from two-cell embryos (75.0%) was similar to that of nonfrozen blastomeres (76.0%). When eight pairs of blastocysts that developed from frozen two-cell mouse blastomeres were transferred to pregnant ICR recipients on Day 3, four live singletons were born.Conclusion Thus, the results indicate that two-cell mouse blastomeres can be frozen by the ultrarapid freezing method.     

Mitral valve surgery with surgical embolectomy for mitral valve endocarditis complicated by septic coronary embolism.

Acute myocardial infarction (AMI) complicated by septic coronary embolism due to active infective endocarditis is rare but usually fatal. We report a case of successful mitral valve surgery with surgical embolectomy in a 27-year-old man with an AMI complicated by septic coronary embolism due to mitral valve endocarditis. A chest radiograph revealed cardiomegaly and marked pulmonary edema. A transthoracic echocardiogram disclosed severe mitral regurgitation with highly mobile vegetations and hypokinesia of the left ventricular apex. The electrocardiographic findings of ST segment elevation in leads V2-4 and elevated cardiac enzyme levels were strongly suggestive of an acute anterolateral AMI. Nevertheless, emergent cardiac surgery was needed without selective coronary angiography because of intractable heart failure and life-threatening ventricular tachyarrhythmia requiring cardiopulmonary resuscitation. A total occlusion of the distal left anterior descending artery caused by embolic vegetation and thrombus, which was incidentally detected intraoperatively, was successfully recanalized by surgical embolectomy and thrombectomy using a direct coronary incision. The mitral valve endocarditis was managed with wide debridement and mechanical valve replacement. Three years after the surgery a follow-up echocardiogram showed no abnormalities of the regional wall, motion in the left ventricle and the patient is living an active life without any complications.     

A novel form of self tolerance dictated in the thymus of transgenic mice with autoreactive TCR {alpha} and {beta} chain genes

Transgenic (TG) mice with TCR and ß chain genes froma CD4-dependent auto-l-A^k reactive T cell clone were generated.H-2^k TG mice had a large number of thymic and splenic CD4 Tcells expressing the autoreactive TCR without manifestationof autolmmunlty. The cells were not anergic, as they could respondto autologous antigen presenting cells and antl-TCR antibodiesin vitro to proliferate and to produce interleuklns. Variousdegrees of down-regulation of CD2 and CD44 was observed in TGmice, Indicating the presence of a defective co-stlmulatoryprocess in TG T cells. These features indicate that the selftolerance in autoreactive TCR TG mice is due not to clonal deletionand anergy but to a novel mechanism where T cells cannot sufficientlyrespond to normally existing self ligand in vivo. That suchan in vivo unresponsiveness of autoreactive T cells is dictatedin the thymus during CD4 T cell differentiation as an atypicalform of positive selection of autoreactive T cells was suggestedby the abnormal surface expression of CD69 and HSA.     

Surgical treatment of chronic atrial fibrillation combined with rheumatic mitral valve disease: Effects of the cryo-maze procedure and predictors for late recurrence.

OBJECTIVE: The aim of this study was to evaluate the effects of the modified maze procedure using cryoablation for treating chronic atrial fibrillation (AF) associated with rheumatic mitral valve disease and to assess the risk factors for late failure of sinus rhythm restoration. METHODS: Between March 2000 and June 2004, 170 consecutive patients, who underwent the modified maze procedure using cryoablation concomitant with mitral valve surgery for atrial fibrillation associated with rheumatic mitral valve disease, were divided into two groups based on the type of right-sided maze: the modified Cox-maze III (CM group, n=93) and modified Kosakai-maze (KM group, n = 77) procedures. The postoperative and mid-term follow-up results were analyzed and compared between the two groups. Both univariate and multivariate analyses were used to assess the risk factors for late recurrence of atrial fibrillation. RESULTS: There were three in-hospital deaths, including two in the CM group (2.2%) and one in the KM group (1.3%), and there were no significant differences in the incidence of postoperative complications between the two groups. The cardiopulmonary bypass and aortic cross-clamp times were significantly shorter in the KM group than the CM group. At discharge, the sinus rhythm rate was 70% in the CM group and 74% in the KM group (p = 0.55). Follow-up was completed in 97% of the patients, with a mean time of 26.6+/-15.2 months. At the latest follow-up, one death occurred in the CM group (0.6%). Sinus rhythm was documented in 141 (84%) out of all the patients, including 76 (84%) in the CM group and 65 (86%) in the KM group (p=0.72). The 4-year actuarial survival free from stroke was 90.3+/-5.9% for the CM group and 96.4+/-3.5% for the KM group (p = 0.68), and 4-year event-free survival was 81.2+/-7.4% for the CM group and 96.4+/-3.5% for the KM group (p = 0.078). Using a multivariate analysis, a left atrial dimension >65 mm (p = 0.011) and repair for rheumatic mitral valve disease (p = 0.038) were independent risk factors for a late recurrence of AF. CONCLUSIONS: The modified maze procedures using cryoablation are safe and effective in treating chronic atrial fibrillation associated with rheumatic mitral valve disease.     

Epigenetic signatures and temporal expression of lineage-specific genes in hESCs during differentiation to hepatocytes in vitro

Embryonic stem cells (ESCs) maintain unique epigenetic states to maintain their pluripotency. Differentiation of ESCs into specialized cell types requires changes in these epigenetic states. However, the dynamics of epigenetic marks found in hESCs during differentiation are poorly understood. Here, we report the variation in the dynamics of epigenetic modifications associated with the expression of lineage-specific genes during differentiation of hESCs to hepatocytes in vitro. The promoter regions of pluripotency marker genes characterized by permissive histone marks such as trimethylation of H3 at lysine 4 (H3K4me3) and acetylation of H3 at lysine 9 (H3K9ac) in hESCs were instead enriched with repressive histone marks such as dimethylation of H3 at lysine 9 (H3K9me2), trimethylation of H3 at lysine 9 (H3K9me3) and trimethylation of H3 at lysine 27 (H3K27me3) during differentiation to hepatocytes. Interestingly, expression of definitive endoderm marker genes containing bivalent and non-bivalent domains may be modulated by a marked reduction in H3K27me3 and a significant enhancement of permissive marks such as H3K4me3 and H3K9ac during hESC differentiation. Expression of hepatocyte marker genes regulated by histone modifications was similar to that of pluripotency marker genes. Our findings provide insight into the epigenetic mechanisms regulating expression of developmental genes. Of particular interest, they may be differentially regulated either in a bivalent or non-bivalent domain manner during hESC differentiation.     

Neutrophil-to-Lymphocyte Ratio at Emergency Room Predicts Mechanical Complications of ST-segment Elevation Myocardial Infarction

BackgroundThe neutrophil-to-lymphocyte ratio (NLR) has been proven to be a reliable inflammatory marker. A recent study reported that elevated NLR is associated with adverse cardiovascular events in patients with ST-segment elevation myocardial infarction (STEMI). We investigated whether NLR at emergency room (ER) is associated with mechanical complications of STEMI undergoing primary percutaneous coronary intervention (PCI).MethodsA total of 744 patients with STEMI who underwent successful primary PCI from 2009 to 2018 were enrolled in this study. Total and differential leukocyte counts were measured at ER. The NLR was calculated as the ratio of neutrophil count to lymphocyte count. Patients were divided into tertiles according to NLR. Mechanical complications of STEMI were defined by STEMI combined with sudden cardiac arrest, stent thrombosis, pericardial effusion, post myocardial infarction (MI) pericarditis, and post MI ventricular septal rupture, free-wall rupture, left ventricular thrombus, and acute mitral regurgitation during hospitalization.ResultsPatients in the high NLR group (> 4.90) had higher risk of mechanical complications of STEMI (P = 0.001) compared with those in the low and intermediate groups (13% vs. 13% vs. 23%). On multivariable analysis, NLR remained an independent predictor for mechanical complications of STEMI (RR = 1.947, 95% CI = 1.136–3.339, P = 0.015) along with symptom-to balloon time (P = 0.002) and left ventricular dysfunction (P < 0.001).ConclusionNLR at ER is an independent predictor of mechanical complications of STEMI undergoing primary PCI. STEMI patients with high NLR are at increased risk for complications during hospitalization, therefore, needs more intensive treatment after PCI.