Catechol-O-methyltransferase (COMT) Val158Met Polymorphism and Susceptibility to Alcohol Dependence

Catechol-O-methyl transferase (COMT) enzyme catalyzes the metabolism of dopamine and other catechols in the brain. Several articles investigated catechol-O-methyltransferase (COMT) Val158Met polymorphism as risk factor for alcohol dependence (AD) but the results were inconclusive. The aim of present meta-analysis was to evaluate the association of Val158Met (COMT) polymorphism with AD. Authors performed keyword search of the 4 electronic databases—Pubmed, Google Scholar, Springer Link and Science Direct databases up to December 31, 2019. Total eighteen studies that investigated the association of Val158Met polymorphism with AD were retrieved. The pooled results from the meta-analysis (2278 AD cases and 3717 healthy controls) did not show association with AD using all 5 genetic models (allele contrast model: OR = 1.02, 95% CI = 0.90–1.14, p = 0.03; homozygote model: OR = 1.06, 95% CI = 0.81–1.38, p = 0.69; dominant model: OR = 0.99, 95% CI = 0.85–1.14, p = 0.87; co-dominant model: OR = 0.97, 95% CI = 0.86–1.11, p = 0.71; recessive model: OR = 1.05;95% CI = 0.85–1.29, p = 0.61). Results of subgroup analysis showed that Val158Met is not risk for AD in Asian and Caucasian population. In conclusion, COMT Val158Met is not a risk factor for alcohol dependence.     

Hyperinsulinemia and Hypoadiponectinemia are Associated with Increased Risk for Occurrence of Ovarian Cancer in Non-diabetic Women of North Indian Population

Ovarian cancer has been emerged as a most common and lethal gynecological malignancy in India. High serum insulin and low adiponectin have been associated with increased risk of ovarian cancer. But their role in development of ovarian cancer is conflicting and little evidence is available. We aimed to evaluate blood levels of insulin and adiponectin in epithelial ovarian cancer (EOC) patients and their association with the risk to develop EOC. The study included following three groups; Group 1: fifty cases of cytohistopathologically confirmed cases of EOC, Group 2: fifty age matched cases of benign ovarian conditions and Group 3: fifty ages matched healthy controls with no evidence of any benign or malignant ovarian pathology as ruled out by clinical examination and relevant investigations. Cytohistopathologically confirmed and newly diagnosed cases of EOC and benign ovarian cancer were included in this study. The median value of fasting serum insulin was significantly high (15.0 µlU/ml, P = 0.02) and adiponectin were significantly low (5.1 µg/ml, P < 0.001) in ovarian cancer patients compared to benign ovarian tumors and healthy controls group. A significant increase risk of ovarian cancer was found in high tertile (≥ 18.7 µlU/ml) of serum insulin level (OR = 2.7; 95% CI = 1.00–6.67, P = 0.04) and lower tertile (≤ 5.45 µg/ml) of adiponectin level (OR = 3.2; 95% CI = 1.10–9.71, P = 0.03). High serum insulin level and low adiponectin levels were significantly associated with increased risk for development of ovarian cancer.     

Genetic Polymorphism of Angiotensin II Type 1 Receptors and Their Effect on the Clinical Outcome of Captopril Treatment in Arab Iraqi Patients with Acute Coronary Syndrome (Mid Euphrates)

Genetic variation in the angiotensin II type 1 receptor (AT1R) has an important effect on the outcome of acute coronary syndrome (ACS) initiated treatment with captopril. This study aims to investigate the impact of genetic polymorphism of AT1R (rs5186 and rs275651) on the ACS outcome in Iraqi patients treated with captopril. A total of 250 Iraqi individuals with ACS were included in this case—control study and they were divided into two study groups; Study group 1 included 125 participants who were prescribed captopril, 25 mg twice daily and study group 2 included 125 participants who received no captopril as part of their ACS treatment (control study). The AT1R gene (rs5186) CC genotype was found to be associated with ST-elevation myocardial infarction (STEMI) (Odd’s ratio (O.R) = 1.2, P = 0.7), while AC was associated with Non-ST-elevation myocardial infarction (NSTEMI) and unstable angina (UA) (O.R = 1.2, P = 0.8). AC genotype is more prone to have Percutaneous coronary intervention (PCI) after ACS attack (O.R = 1.2, P = 0.6). CC genotype had a risk to get less improvement (O.R = 1.6, P = 0.5), so might require higher doses of captopril during acute coronary insult. The AT1R gene (rs275651) AA genotype was associated with UA (O.R = 1.3, P = 0.9). AA and AT genotypes were more prone to have PCI after ACS attack (O.R = 3.9 P = 0.2, O.R = 3.5, P = 0.3 respectively) and thus requiring higher doses of captopril. We conclude that the AT1R rs5186, rs275651 genetic polymorphisms might partially affect the clinical outcome of ACS patients treated with captopril and might have captopril resistance which requires higher doses.     

Synergetic Interaction of HLA-DRB1*07 Allele and TNF-Alpha − 863 C/A Single Nucleotide Polymorphism in the Susceptibility to Systemic Lupus Erythematosus

Systemic lupus erythematosus (SLE) is an inflammatory autoimmune disease which is characterized by dysregulation of various cytokines propagating the inflammatory processes that is responsible for tissue damage. Tumor necrosis factor alpha (TNF-α) is one of the most important immunoregulatory cytokines that has been implicated in the different autoimmune diseases including SLE. Two hundred and two patients with SLE and 318 controls were included in the study. The TNF-α gene promoter region (from − 250 to − 1000 base pairs) was analyzed by direct Sanger’s DNA sequencing method to find promoter variants associated with South Indian SLE patients. We have analyzed six TNF-α genetic polymorphisms including, − 863C/A (rs1800630), − 857C/T (rs1799724), − 806C/T (rs4248158), − 646G/A (rs4248160), − 572A/C (rs4248161) and − 308G/A (rs1800629) in both SLE patients and controls. We did not find association of TNF-α gene promoter SNPs with SLE patients. However, the − 863A (rs1800630) allele showed association with lupus nephritis phenotype in patients with SLE (OR: 1.62, 95%CI 1.04–2.53, P = 0.034). We found serum TNF-α level was significantly elevated in SLE cases as compared to control and found no association with any of the polymorphisms. The haplotype analysis revealed a significant protective association between the wild TNF-α alleles at positions − 863C, − 857C, − 806C, − 646G, − 572A and − 308G (CCCGAG) haplotype with lupus nephritis phenotype (OR 0.53, 95% CI 0.35–0.82, P = 0.004). Additionally, the TNF-α − 863 C/A (rs1800630) polymorphism and HLA-DRB1*07 haplotype showed significant differences between SLE patients and controls (OR 4.79, 95% CI 1.73–13.29, P = 0.0009). In conclusion, TNF-α − 863A allele (rs1800630) polymorphism is associated with increased risk of nephritis in South Indian SLE patients. We also found an interaction between HLA-DRB1*07 allele with TNF-α − 863 C/A promoter polymorphism giving supportive evidence for the tight linkage disequilibrium between TNF-α promoter SNPs and MHC class II DRB1 alleles.     

Genetic Variant XRCC1 rs1799782 (C194T) and Risk of Cancer Susceptibility in Indian Population: A Meta-analysis of Case–Control Studies

X-ray repair cross-complementing group 1 (XRCC1) plays a key role in the base excision repair pathway, as a scaffold protein that brings together proteins of the DNA repair complex. Several studies have reported contradictory results for XRCC1 exon 6 C>T (rs1799782) gene polymorphism and cancer risk in Indian population has provided inconsistent results. Therefore, we have performed this meta-analysis to evaluate the relationship between XRCC1 exon 6 C>T gene polymorphism and risk of cancer by published studies. We searched PubMed and Google scholar web databases to cover all studies published on association between XRCC1 exon 6 C>T gene polymorphism and cancer risk. The meta-analysis was carried out and pooled odds ratios (ORs) and 95% confidence intervals (95% CIs) were used to appraise the strength of association. In order to derive a more precise estimation of the association, A total of 3197 confirmed cancer cases and 3819 controls were included from eligible seventeen case-controls studies. Results from overall pooled analysis demonstrated suggested that that variant allele (T vs. C: OR 1.301, 95% CI 1.003–1.688, p = 0.047) was associated with the risk of overall cancer. Other genetic models; heterozygous (TC vs. CC: OR 1.108, 95% CI 0.827–1.485, p = 0.491), homozygous (TT vs. CC: OR 1.479, 95% CI 0.877–2.493, p = 0.142), dominant (TT+TC vs. CC: OR 1.228, 95% CI 0.899–1.677, p = 0.196) and recessive (TT vs. TC+CC: OR 1.436, 95% CI 0.970–2.125, p = 0.071) did not reveal statistical association. Publication bias observation was also considered and none was detected during the analysis. The present meta-analysis suggested that the variant allele T of XRCC1 exon 6 gene polymorphism was associated with the risk of cancer. It is therefore pertinent to confirm this finding in a large sample size to divulge the mechanism of this polymorphism and cancer risk in Indian population.     

Genetic Association of rs2237572 Cyclin-Dependent Kinase 6 Gene with Breast Cancer in Iraq

This case–control study is aimed to evaluate serum concentration of Cyclin-Dependent Kinase 6 (CDK6) and the genetic association between rs2237572 CDK6 gene and breast cancer (BC) in Iraq. To attain this goal, 80 patients with BC as cases and 80 healthy individuals as controls were included. Further, BC patients were sorted according to the molecular classification into four subtypes of Luminal A, Luminal B, Her2/neu enriched and TPN. Serum concentration of CDK6 enzyme, allelic and genotypic frequencies of rs2237572 CDK6, and the occurrence of BC phenotype and its subtypes in the studied population were investigated. ELISA technique was used to perform the biochemical testing, while the molecular analysis was achieved by real-time PCR, high resolution melting analysis, conventional PCR, as well as sequencing analysis. The results revealed no significant difference in serum concentration of CDK6 enzyme between patients and healthy controls (p > 0.05). Also, no significant differences were shown between BC patients subtypes (p > 0.05). The rs2237572 CDK6 genotypes were associated with the BC and affirmed that allele C was inherited as a recessive risk factor. Moreover, a highly significant difference between patients’ subtypes in the genotypic frequency of rs2237572 (p < 0.01) was noted. Furthermore, the association of rs2237572 genotypes and CDK6 serum concentration in BC patients showed a considered significant difference between C/C and T/T, C/C and T/C and the CDK6 level (p < 0.05). Nevertheless, T/T and T/C did not show any significant difference with the CDK6 level. Hence, it was concluded that the rs2237572 of CDK6 gene is significantly correlated with BC.     

Relationship of Brain-Derived Neurotrophic Factor with Interleukin-23, Testosterone and Disease Severity in Schizophrenia

Hormonal imbalance, inflammation and alteration in synaptic plasticity are reported to play a crucial role in the pathogenesis of schizophrenia. The objective of the study was to assess the serum levels of brain derived neurotrophic factor (BDNF) and its association with interleukin-23 (IL-23), testosterone and disease severity in schizophrenia. 40 cases and 40 controls were included in the study. Serum levels of BDNF, IL-23 and testosterone were estimated in all the subjects. Disease severity was assessed using Positive and Negative Syndrome Scale (PANSS). The study was designed in Tertiary care hospital, South India. The results were compared between two groups using Mann–Whitney U test. Spearman Correlation analysis was used to assess the association between biochemical parameters and PANSS. Interleukin-23 and testosterone were significantly increased and BDNF was significantly reduced in schizophrenia cases when compared with controls. BDNF was negatively correlated with IL-23 (r = − 400, p = 0.011), positive symptom subscale (r = − 0.393, p = 0.012), general psychopathology score subscale (r = − 407, p = 0.009) and total symptom subscale (r = − 404, p = 0.010). There was no significant association of IL-23 and testosterone with disease severity in schizophrenia cases. BDNF was reduced in schizophrenia cases and negatively associated with interleukin-23 and disease severity scores.     

C677T MTHFR Gene Polymorphism is Contributing Factor in Development of Renal Impairment in Young Hypertensive Patients

Homocysteine concentration affected by the activities of the enzymes methylene tetra-hyrdofolate reductase (MTHFR). Polymorphisms in MTHFR gene associated with an impairment of MTHFR activity. Hyperhomocysteinemia is a result of single nucleotide polymorphisms (SNPs) in MTHFR 677 C>T that can cause homocysteine levels in the blood to increase. The purpose of this study is to investigate the relationships between MTHFR C677T (rs1801133) gene polymorphism, changes in homocysteine concentrations and progress of renal impairment in young adult hypertensive patients. Two hundred young hypertensive patients (age 21–24 years) were involved in this study; they were classified into patients with and without renal impairment in addition to 200 age and sex matched healthy controls. All participants were submitted to laboratory investigations as assay of MTHFR gene polymorphism C677T (rs1801133) by PCR/RFLP, determination of lipid profile, homocysteine and folic acid concentrations in addition to urinary albumin creatinine ratio (UACR). The levels of both homocysteine and UACR in the TT genotype patients were higher than those in the CC genotype group. Individuals who carry the T allele were more risky to hypertension and progress to early renal impairment in young age compared with those carrying the C allele [OR 2.02 (1.33–3.08), P < 0.001]. Genetic variants of C677T MTHFR gene and hyperhomocysteinemia may be responsible for rapid progress of renal impairment in Egyptian young age hypertensive patients. TT genotype or T allele may be considered as a predisposing factor for both elevated Hcy levels and the development of renal impairment. This study believed that lowering of homocysteine level can reduce renal impairment of hypertensive patients.     

Correlating the electronic structures of metallic/semiconducting MoTe2 interface to its atomic structures

Contact interface properties are important in determining the performances of devices that are based on atomically thin two-dimensional (2D) materials, especially for those with short channels. Understanding the contact interface is therefore important to design better devices. Herein, we use scanning transmission electron microscopy, electron energy loss spectroscopy, and first-principles calculations to reveal the electronic structures within the metallic (1T^′)-semiconducting (2H) MoTe₂ coplanar phase boundary across a wide spectral range and correlate its properties to atomic structures. We find that the 2H-MoTe₂ excitonic peaks cross the phase boundary into the 1T^′ phase within a range of approximately 150 nm. The 1T^′-MoTe₂ crystal field can penetrate the boundary and extend into the 2H phase by approximately two unit-cells. The plasmonic oscillations exhibit strong angle dependence, that is a red-shift of π+σ (approximately 0.3–1.2 eV) occurs within 4 nm at 1T^′/2H-MoTe₂ boundaries with large tilt angles, but there is no shift at zero-tilted boundaries. These atomic-scale measurements reveal the structure–property relationships of the 1T^′/2H-MoTe₂ boundary, providing useful information for phase boundary engineering and device development based on 2D materials.     

Adiponectin Gene Polymorphism and its Association with Type 2 Diabetes Mellitus

Mutations in different regions of adiponectin gene have been reported to be associated with obesity, atherosclerosis and type 2 diabetes mellitus. The present study was aimed to investigate the association among SNP 45 T > G of adiponectin gene and type 2 diabetes in South Indian population. 75 clinically diagnosed case of type 2 diabetes were studied and compared with 75 apparently healthy controls. The genotype frequency of SNP45 T > G in exon 2 of adiponectin gene was determined by PCR based restriction enzyme analysis using the restriction enzyme SmaI. (recognition site: CCC↓GGG). Three kind of genotypes: wild type TT (470 bp), heterozygous type TG (470 bp, 336 bp, 134 bp) and homozygote mutant type GG (336 bp, 134 bp) were studied. A positive association has been found between SNP45 T > G and type 2 diabetes in the study population (P = 0.010, OR = 3.797, 95% CI = 1.312–10.983). Therefore, SNP45T > G in adiponectin gene may be one of the risk factors for type 2 diabetes.     

Blood viscoelasticity measurement using steady and transient flow controls of blood in a microfluidic analogue of Wheastone-bridge channel

Accurate measurement of blood viscoelasticity including viscosity and elasticity is essential in estimating blood flows in arteries, arterials, and capillaries and in investigating sub-lethal damage of RBCs. Furthermore, the blood viscoelasticity could be clinically used as key indices in monitoring patients with cardiovascular diseases. In this study, we propose a new method to simultaneously measure the viscosity and elasticity of blood by simply controlling the steady and transient blood flows in a microfluidic analogue of Wheastone-bridge channel, without fully integrated sensors and labelling operations. The microfluidic device is designed to have two inlets and outlets, two side channels, and one bridge channel connecting the two side channels. Blood and PBS solution are simultaneously delivered into the microfluidic device as test fluid and reference fluid, respectively. Using a fluidic-circuit model for the microfluidic device, the analytical formula is derived by applying the linear viscoelasticity model for rheological representation of blood. First, in the steady blood flow, the relationship between the viscosity of blood and that of PBS solution (μ_Blood/μ_PBS) is obtained by monitoring the reverse flows in the bridge channel at a specific flow-rate rate (Q_PBS^SS/Q_Blood^L). Next, in the transient blood flow, a sudden increase in the blood flow-rate induces the transient behaviors of the blood flow in the bridge channel. Here, the elasticity (or characteristic time) of blood can be quantitatively measured by analyzing the dynamic movement of blood in the bridge channel. The regression formula (A_Blood (t) = A_α + A_β exp [−(t − t₀)/λ_Blood]) is selected based on the pressure difference (ΔP = P_A − P_B) at each junction (A, B) of both side channels. The characteristic time of blood (λ_Blood) is measured by analyzing the area (A_Blood) filled with blood in the bridge channel by selecting an appropriate detection window in the microscopic images captured by a high-speed camera (frame rate = 200 Hz, total measurement time = 7 s). The elasticity of blood (G_Blood) is identified using the relationship between the characteristic time and the viscosity of blood. For practical demonstrations, the proposed method is successfully applied to evaluate the variations in viscosity and elasticity of various blood samples: (a) various hematocrits form 20% to 50%, (b) thermal-induced treatment (50 °C for 30 min), (c) flow-induced shear stress (53 ± 0.5 mL/h for 120 min), and (d) normal rat versus spontaneously hypertensive rat. Based on these experimental demonstrations, the proposed method can be effectively used to monitor variations in viscosity and elasticity of bloods, even with the absence of fully integrated sensors, tedious labeling and calibrations.     

Two-dimensional ferromagnetic superlattices

Mechanically exfoliated two-dimensional ferromagnetic materials (2D FMs) possess long-range ferromagnetic order and topologically nontrivial skyrmions in few layers. However, because of the dimensionality effect, such few-layer systems usually exhibit much lower Curie temperature (T_C) compared to their bulk counterparts. It is therefore of great interest to explore effective approaches to enhance their T_C, particularly in wafer-scale for practical applications. Here, we report an interfacial proximity-induced high-T_C 2D FM Fe₃GeTe₂ (FGT) via A-type antiferromagnetic material CrSb (CS) which strongly couples to FGT. A superlattice structure of (FGT/CS)_n, where n stands for the period of FGT/CS heterostructure, has been successfully produced with sharp interfaces by molecular-beam epitaxy on 2-inch wafers. By performing elemental specific X-ray magnetic circular dichroism (XMCD) measurements, we have unequivocally discovered that T_C of 4-layer Fe₃GeTe₂ can be significantly enhanced from 140 K to 230 K because of the interfacial ferromagnetic coupling. Meanwhile, an inverse proximity effect occurs in the FGT/CS interface, driving the interfacial antiferromagnetic CrSb into a ferrimagnetic state as evidenced by double-switching behavior in hysteresis loops and the XMCD spectra. Density functional theory calculations show that the Fe-Te/Cr-Sb interface is strongly FM coupled and doping of the spin-polarized electrons by the interfacial Cr layer gives rise to the T_C enhancement of the Fe₃GeTe₂ films, in accordance with our XMCD measurements. Strikingly, by introducing rich Fe in a 4-layer FGT/CS superlattice, T_C can be further enhanced to near room temperature. Our results provide a feasible approach for enhancing the magnetic order of few-layer 2D FMs in wafer-scale and render opportunities for realizing realistic ultra-thin spintronic devices.     

Influence of Altered Hormonal Status on Platelet 5-HT and MAO-B Activity in Cigarette Smokers

The present study was designed to understand the cigarette smoking-induced alterations in hormones and the resulting changes in platelet serotonin (5-hydroxytryptamine, 5-HT) and monoamine oxidase (MAO-B) activity in chronic smokers. Human male volunteers aged 35 ± 8 years, were divided into two groups, namely controls and smokers (12 ± 2 cigarettes per day for 7–10 years). Results showed that cigarette smoking significantly (p < 0.05) elevated plasma triiodothyronine (T₃), cortisol and testosterone levels with significant (p < 0.05) reduction in plasma tryptophan and thyroxin (T₄). Moreover, smokers showed reduced platelet 5-HT levels and MAO-B activity. In smokers, plasma cortisol was negatively correlated with tryptophan (r = −0.386), platelet MAO-B (r = −0.264), and 5-HT (r = −0.671), and positively correlated with testosterone (r = 0.428). However, testosterone was negatively correlated with platelet MAO-B (r = −0.315), and 5-HT (r = −.419) in smokers. Further, smokers plasma T₃ levels were negatively correlated with platelet MAO-B (r = −0.398), and 5-HT (r = −0.541), whereas T₄ levels were positively correlated with platelet MAO-B (r = 0.369), and 5-HT (r = 0.454). In conclusion, our study showed that altered testosterone and cortisol levels may aggravate behavior, mood disturbances and symptoms of depression by decreasing platelet 5-HT and MAO-B activity in smokers.     

Micromixing visualization and quantification in a microscale multi-inlet vortex nanoprecipitation reactor using confocal-based reactive micro laser-induced fluorescence

A technique for visualizing and quantifying reactive mixing for laminar and turbulent flow in a microscale chemical reactor using confocal-based microscopic laser induced fluorescence (confocal μ-LIF) was demonstrated in a microscale multi-inlet vortex nanoprecipitation reactor. Unlike passive scalar μ-LIF, the reactive μ-LIF technique is able to visualize and quantify micromixing effects. The confocal imaging results indicated that the flow in the reactor was laminar and steady for inlet Reynolds numbers of 10, 53, and 93. Mixing and reaction were incomplete at each of these Reynolds numbers. The results also suggested that although mixing by diffusion was enhanced near the midplane of the reactor at Re_j = 53 and 93 due to very thin bands of acidic and basic fluid forming as the fluid spiraled towards the center of the reactor, near the top, and bottom walls of the reactor, the lower velocities due to fluid friction with the walls hindered the formation of these thin bands, and, thus, resulted in large regions of unmixed and unreacted fluid. At Re_j = 240, the flow was turbulent and unsteady. The mixing and reaction processes were still found to be incomplete even at this highest Reynolds number. At the reactor midplane, the flow images at Re_j = 240 showed unmixed base fluid near the center of the reactor, suggesting that just as in the Re_j = 53 and 93 cases, lower velocities near the top and bottom walls of the reactor hinder the mixing and rection of the acidic and basic streams. Ensemble averages of line-scan profiles for the Re_j = 240 were then calculated to provide statistical quantification of the microscale mixing in the reactor. These results further demonstrate that even at this highest Reynolds number investigated, mixing and reaction are incomplete. Visualization and quantification of micromixing using this reactive μ-LIF technique can prove useful in the validation of computational fluid dynamics models of micromixing within microscale chemical reactors.     

Influence of buoyancy-driven flow on mass transfer in a two-stream microfluidic channel: Introduction of cryoprotective agents into cell suspensions

A variety of methods have been used to introduce chemicals into a stream or to mix two or more streams of different compositions using microfluidic devices. In the following paper, the introduction of cryoprotective agents (CPAs) used during cryopreservation of cells in order to protect them from freezing injuries and increase viability post thaw is described. Dimethylsulphoxide (DMSO) is the most commonly used CPA. We aim to optimize the operating conditions of a two-stream microfluidic device to introduce a 10% vol/vol solution of DMSO into a cell suspension. Transport behavior of DMSO between two streams in the device has been experimentally characterized for a spectrum of flow conditions (0.7 < Re < 10), varying initial donor stream concentrations, (1% vol/vol < C_o < 15% vol/vol) and different flow rate fractions (0.23 < f_q < 0.77). The outlet cell stream concentration is analyzed for two different flow configurations: one with the cell stream flowing on top of the DMSO-rich donor stream, and the other with the cell stream flowing beneath the heavy DMSO-laden stream. We establish a transition from a diffusive mode of mass transfer to gravity-influenced convective currents for Atwood numbers (A_t) in the range of (1.7 × 10⁻³ < A_t < 3.1 × 10⁻³) for the latter configuration. Flow visualization with cells further our understanding of the effect of A_t on the nature of mass transport. Cell motion studies performed with Jurkat cells confirm a high cell recovery from the device while underscoring the need to collect both the streams at the outlet of the device and suggesting flow conditions that will help us achieve the target DMSO outlet concentration for clinical scale flow rates of the cell suspension.     

Microwells support high-resolution time-lapse imaging and development of preimplanted mouse embryos

A vital aspect affecting the success rate of in vitro fertilization is the culture environment of the embryo. However, what is not yet comprehensively understood is the affect the biochemical, physical, and genetic requirements have over the dynamic development of human or mouse preimplantation embryos. The conventional microdrop technique often cultures embryos in groups, which limits the investigation of the microenvironment of embryos. We report an open microwell platform, which enables micropipette manipulation and culture of embryos in defined sub-microliter volumes without valves. The fluidic environment of each microwell is secluded from others by layering oil on top, allowing for non-invasive, high-resolution time-lapse microscopy, and data collection from each individual embryo without confounding factors. We have successfully cultured mouse embryos from the two-cell stage to completely hatched blastocysts inside microwells with an 89% success rate (n = 64), which is comparable to the success rate of the contemporary practice. Development timings of mouse embryos that developed into blastocysts are statistically different to those of embryos that failed to form blastocysts (p–value < 10⁻¹⁰, two-tailed Student''s t-test) and are robust indicators of the competence of the embryo to form a blastocyst in vitro with 94% sensitivity and 100% specificity. Embryos at the cleavage- or blastocyst-stage following the normal development timings were selected and transferred to the uteri of surrogate female mice. Fifteen of twenty-two (68%) blastocysts and four of ten (40%) embryos successfully developed into normal baby mice following embryo transfer. This microwell platform, which supports the development of preimplanted embryos and is low-cost, easy to fabricate and operate, we believe, opens opportunities for a wide range of applications in reproductive medicine and cell biology.