Engagement of Robo1 by Slit2 induces formation of a trimeric complex consisting of Src-Robo1-E-cadherin for E-cadherin phosphorylation and epithelial-mesenchymal transition.

Loss of E-cadherin elicits epithelial-mesenchymal transition (EMT). While both the Src family of membrane-associated non-receptor tyrosine kinases (SFKs) and Slit2 binding to Roundabout 1 (Robo1) have been shown to induce E-cadherin repression and EMT, whether these two signaling pathways are mechanistically coupled remains unknown in epithelial cells. Here we found that Slit2 and Robo1 overexpression activated Src kinases for tyrosine phosphorylation, degradation of E-cadherin and induction of EMT. Specific blockade of Slit2 binding to Robo1 inactivated Src, prevented E-cadherin phosphorylation and EMT induction. Biochemically, the cytoplasmic CC3 motif of Robo1 (CC3) bound directly to the SH2 and 3 domains of c-Src and the cytoplasmic domains of E-cadherin. Slit2 induced Robo1 association with endogenous c-Src and E-cadherin, whereas ectopic expression of CC3 dissociated this protein complex in colorectal epithelial cells. These results indicate that Slit2 not only induces Robo1 binding to Src, but also recruits Src to E-cadherin for tyrosine phosphorylation of E-cadherin, leading to E-cadherin degradation and EMT induction in colorectal epithelial cells.

Eukaryotic translation initiation factor 5A2 promotes metabolic reprogramming in hepatocellular carcinoma cells.

Reprogramming of intracellular metabolism is common in liver cancer cells. Understanding the mechanisms of cell metabolic reprogramming may present a new basis for liver cancer treatment. In our previous study, we reported that a novel oncogene eukaryotic translation initiation factor 5A2 (EIF5A2) promotes tumorigenesis under hypoxic condition. Here, we aim to investigate the role of EIF5A2 in cell metabolic reprogramming during hepatocellular carcinoma (HCC) development. In this study, we reported that the messenger RNA (mRNA) level of EIF5A2 was upregulated in 59 of 105 (56.2%) HCC clinical samples (P = 0.015), and EIF5A2 overexpression was significantly associated with shorter survival time of patients with HCC (P = 0.021). Ectopic expression of EIF5A2 in HCC cell lines significantly promoted cell growth and accelerated glucose utilization and lipogenesis rates. The high rates of glucose uptake and lactate secretion conferred by EIF5A2 revealed an abnormal activity of aerobic glycolysis in HCC cells. Several key enzymes involved in glycolysis including glucose transporter type 1 and 2, hexokinase 2, phosphofructokinase liver type, glyceraldehyde 3-phosphate dehydrogenase, pyruvate kinase M2 isoform, phosphoglycerate mutase 1 and lactate dehydrogenase A were upregulated by overexpression of EIF5A2. Moreover, EIF5A2 showed positive correlations with FASN and ACSS2, two key enzymes involved in the fatty acid de novo biosynthetic pathway, at both protein and mRNA levels in HCC. These results indicated that EIF5A2 may regulate fatty acid de novo biosynthesis by increasing the uptake of acetate. In conclusion, our findings demonstrate that EIF5A2 has a critical role in HCC cell metabolic reprogramming and may serve as a prominent novel therapeutic target for liver cancer treatment.

Toxicity and accumulation of zinc pyrithione in the liver and kidneys of Carassius auratus gibelio: association with P-glycoprotein expression.

Zinc pyrithione (ZPT) is a broad-spectrum antibacterial and antifungal agent; therefore, it is widely used in industry and civilian life. It is discharged into the aquatic environment with industrial and civilian waste water. Carassius sp. is one of the most widely distributed and farmed fish in China. The effects of aquatic ZPT on Carassius sp. remain unknown. In this study, we determined the acute toxicity of ZPT on Carassius sp. The results showed that the median lethal concentration (LC50 96 h) of ZPT on Carassius sp. cultivated in freshwater or water with 1.5 or 3 ‰ salinity was 0.163, 0.126, and 0.113 mg/L, respectively. ZPT has a higher affinity to the liver than the kidney, with a prolonged tissue residual time. P-glycoprotein (P-gp), an ATP-binding cassette transporter, was found to be induced in the liver and kidney tissues of these Carassius spp. after ZPT treatment, based on the determination of its mRNA and protein levels by quantitative real-time reverse transcription polymerase chain reaction and immunohistochemistry, respectively. The ZPT accumulation and magnitude of P-gp induction were also affected by the salinity of the cultivation water. These results suggest that aquatic ZPT is potentially toxic to Carassius sp. We speculate that P-gp induction may play a protective role for Carassius sp. Our findings provide a basis for assessing the potential risk of ZPT to aquatic animals including Carassius sp.

[Association between CMTM5 gene rs723840 single nucleotide polymorphism and high on asprin platelet reactivity].

OBJECTIVE: To elucidate the correlation between the single nucleotide polymorphism of CKLF-like MARVEL transmembrane member 5 (CMTM5) gene rs723840 and the occurrence of high on aspirin platelet reactivity (HAPR). METHODS: The present study is a case-control study. A total of 210 hospitalized patients in Peking University First Hospital were enrolled. Aspirin response was assessed by 0.5 g/L arachidonic acid (AA)-induced platelet aggregation ratio (PR), and ≥ 3/4 quartile of PR of the population was defined as HAPR. Accordingly all the enrolled 210 coronary artery diseases (CAD) patients were divided into HAPR group and No-HAPR group. The genotypes were determined by polymerase chain reaction (PCR) and sequencing analysis for rs723840 of CMTM5 gene. RESULTS: The genotype frequencies in rs723840 C>T of CMTM5 gene conformed well to the Hardy-Weinberg equilibrium in both HAPR group and No-HAPR group. Between the two groups, the genotypes frequencies in HAPR and No-HAPR groups were 48.4%, 51.6%, 0.0% and 73.7%, 22.9%, 0.034%, respectively (P=0.004). The C, T allele frequencies were significantly different in the two groups (P=0.031,OR=0.501, 95% CI: 0.264-0.947). CONCLUSION: Our study finds a significant correlation between CMTM5 gene rs723840 polymorphism and high on aspirin platelet reactivity.

[Correlation between the level of the urinary 11-dehydrothromboxane B2 and the clinical efficacy of aspirin in patients with type 2 diabetes and coronary artery disease].

OBJECTIVE: To elucidate the correlation between urinary 11-dehydro-thromboxane B2 (11dhTxB2) and clinical efficacy of aspirin treatment in patients with type 2 diabete and coronary artery disease (CAD). METHODS: In this prospective cohort study, 169 aged patients with type 2 diabete accompanying CAD in Peking University First Hospital were enrolled. The level of urinary 11dhTxB2 was detected using enzyme-linked immuno-sorbent assay. Low aspirin response or high on aspirin platelet reactivity (HAPR) was defined as urinary 11dhTxB2>1 500 ng/g. All the included patients were divided into two groups based on the results, HAPR group and No-HAPR group. RESULTS: Baseline urinary 11dhTxB2 of the patients with type 2 diabete accompanying CAD was (3 687±3 052) ng/g, while the urinary 11dhTxB2 was (1 954±859) ng/g in patients after 100 mg/d aspirin treatment (P<0.001). Prevalence of HAPR in patients with type 2 diabete accompanying CAD were 32.5%. Within a mean follow-up time of 12 months, the outcomes occurred more frequently in HAPR group than in No-HAPR group (P<0.05). CONCLUSION: Urinary 11dhTxB2 can be recognized as an effective indicator in evaluating aspirin clinical efficacy of patients with type 2 diabete accompanying CAD.

Correlation between expression of thymidylate synthase and clinical outcome of advanced gastric cancer treated with capecitabine alone chemotherapy.

Thymidylate synthase (TS) is a prognostic marker in various tumors. However, the results of previous investigations in gastric cancer (GC) were controversial. The objective of this article is to investigate whether TS expression is associated with clinical outcome in advanced GC receiving capecitabine alone chemotherapy. The study reviewed 58 cases of advanced GC in patients aged ≥65 years between December 2008 and June 2012. All patients were treated with capecitabine alone chemotherapy. Immunohistochemical staining for TS protein expression was performed. The relationships between TS expression and clinicopathological characteristics (included age, gender, number of metastatic sites, Eastern Cooperative Oncology Group (ECOG) score, differentiation, and lymph node metastatic status), chemotherapy response, progression-free survival (PFS), and overall survival (OS) were evaluated. There was no association between TS expression and age, gender, number of metastatic sites, ECOG score, differentiation, and lymph node metastatic status (P > 0.05). The chemotherapy response rates among patients with low- and high-level expression of TS protein were 52.0 % (13/25) and 21.2 % (7/33), respectively (χ (2) = 5.968, P = 0.015). The median PFS and OS in patients with low-level TS expression were significantly longer than those with high-level TS expression (PFS 8.0 vs 2.8 m, P = 0.001; OS 13.3 vs 7.9 m, P = 0.002, respectively). Multivariate Cox regression analysis revealed that TS expression was independent risk factor for OS (hazard ratio (HR) 0.237; 95 % confidence interval (CI) 0.108 to 0.520; P = 0.000). The present study demonstrates that TS expression is associated with chemotherapy response, PFS, and OS in advanced GC patients treated with capecitabine alone chemotherapy.

[Optimization of fermentation techniques of Massa Medicata Fermentata by response surface methodology].

OBJECTIVE: For the first time, the fermentation conditions of Massa Medicata Fermentata were optimized by response surface methodology (RSM) accroding to amylase enzyme activity. METHODS: Firstly, a series of single factor experiments were done which included the amount of Adzuki beans, fermentation time and fermentation temperature. The Box-Behnken design was used to optimize fermentation conditions. Design-Expert V8.0.6.1 was used to analysis regression model. RESULTS: The optimum fermentation conditions were determined as follows: the amount of nitrogen source 2.6 g/100 g wheat bran and wheat flour, fermentation temperature 32 °C and fermentation time 3 days. By comparing prediction value and experimental value, it was found that relative error of their enzyme activity was less than 5%. CONCLUSION: This model can well forecast the amylase enzyme activity of Massa Medicata Fermentata after fermentation. These parameters obtained by RSM can provide scientific basis for the fermentation technology of Massa Medicata Fermentata, and it has a certain practical meaning.

Enantioselective Formal 1,2-Diamination of Ketenes with Iminosulfinamides: Asymmetric Synthesis of Unnatural α,α-Disubstituted α-Amino Acid Derivatives.

A method was developed for the enantioselective formal 1,2-diamination of disubstituted ketenes using iminosulfinamides as nitrogen sources. The protocol involves the addition of lithium iminosulfinamides to ketenes to form N-iminosulfinyl amide metalloenolates. These metalloenolates then undergo a [2,3]-sigmatropic rearrangement to yield unnatural α,α-disubstituted α-amino acid derivatives with high enantiopurity. The chirality present at the sulfur atom in the iminosulfinamides is effectively transferred to α carbon of the resulting products, facilitating the highly enantioselective amination of ketenes.

Predictive value of positive lymph node ratio in patients with locally advanced gastric remnant cancer.

BACKGROUND: Traditional lymph node stage (N stage) has limitations in advanced gastric remnant cancer (GRC) patients; therefore, establishing a new predictive stage is necessary. AIM: To explore the predictive value of positive lymph node ratio (LNR) according to clinicopathological characteristics and prognosis of locally advanced GRC. METHODS: Seventy-four patients who underwent radical gastrectomy and lymphadenectomy for locally advanced GRC were retrospectively reviewed. The relationship between LNR and clinicopathological characteristics was analyzed. The survival analysis was performed using Kaplan-Meier survival curves and Cox regression model. RESULTS: Number of metastatic LNs, tumor diameter, depth of tumor invasion, Borrmann type, serum tumor biomarkers, and tumor-node-metastasis (TNM) stage were correlated with LNR stage and N stage. Univariate analysis revealed that the factors affecting survival included tumor diameter, anemia, serum tumor biomarkers, vascular or neural invasion, combined resection, LNR stage, N stage, and TNM stage (all P < 0.05). The median survival time for those with LNR0, LNR1, LNR2 and LNR3 stage were 61, 31, 23 and 17 mo, respectively, and the differences were significant (P = 0.000). Anemia, tumor biomarkers and LNR stage were independent prognostic factors for survival in multivariable analysis (all P < 0.05). CONCLUSION: The new LNR stage is uniquely based on number of metastatic LNs, with significant prognostic value for locally advanced GRC, and could better differentiate overall survival, compared with N stage.

Establishment and Characterization of an Epstein-Barr Virus-positive Cell Line from a Non-keratinizing Differentiated Primary Nasopharyngeal Carcinoma.

Nasopharyngeal carcinoma (NPC), a cancer that is etiologically associated with the Epstein-Barr virus (EBV), is endemic in Southern China and Southeast Asia. The scarcity of representative NPC cell lines owing to the frequent loss of EBV episomes following prolonged propagation and compromised authenticity of previous models underscores the critical need for new EBV-positive NPC models. Herein, we describe the establishment of a new EBV-positive NPC cell line, designated NPC268 from a primary non-keratinizing, differentiated NPC tissue. NPC268 can undergo productive lytic reactivation of EBV and is highly tumorigenic in immunodeficient mice. Whole-genome sequencing revealed close similarities with the tissue of origin, including large chromosomal rearrangements, while whole-genome bisulfite sequencing and RNA sequencing demonstrated a hypomethylated genome and enrichment in immune-related pathways, respectively. Drug screening of NPC268 together with six other NPC cell lines using 339 compounds, representing the largest high-throughput drug testing in NPC, revealed biomarkers associated with specific drug classes. NPC268 represents the first and only available EBV-positive non-keratinizing differentiated NPC model, and extensive genomic, methylomic, transcriptomic, and drug response data should facilitate research in EBV and NPC, where current models are limited. SIGNIFICANCE: NPC268 is the first and only EBV-positive cell line derived from a primary non-keratinizing, differentiated nasopharyngeal carcinoma, an understudied but important subtype in Southeast Asian countries. This model adds to the limited number of authentic EBV-positive lines globally that will facilitate mechanistic studies and drug development for NPC.

Lead helix winding tricuspid chordae tendineae: A case report.

BACKGROUND: As left bundle branch pacing (LBBP) is more like physiological pacing, LBBP has emerged as a novel pacing strategy that uses the native conduction system to improve ventricular synchronization with stable pacing parameters. LBBP has been revealed associated with a significantly reduced risk of new-onset atrial fibrillation and heart failure compared with conventional permanent pacemaker implantation. CASE SUMMARY: A 64-year-old man was admitted with a 24-h history of chest distress and shortness of breath, which continued unabated. The patient had no symptoms of chest pain, dizziness, syncope, nausea nor vomiting. There were no abnormalities found in routine examinations after admission. Twelve-lead electrocardiogram presented a result of 2:1 atrioventricular block. Coronary angiography was performed the next day and no abnormality was found. Finally, the patient agreed to received LBBP and signed the informed consent. During the process of withdrawing the Medtronic Model 3830 lead into sheath, we found the lead helix was wrapped around the chordae tendineae of the septal valve of tricuspid. Attempts to rotate the 3830 lead failed to release the lead helix from the chordae tendineae, and ultimately we used radiofrequency ablation to ablate the wrapped chordae tendineae. CONCLUSION: Radiofrequency ablation effectively solved this problem without complications. It is an effective and reliable method to resolve lead winding chordae.

Enantioselective Formation of α-Amino Acid Derivatives via [2,3]-Sigmatropic Rearrangement of N-Acyl Iminosulfinamides.

An aza-variation on [2,3]-sigmatropic rearrangement of allylic sulfimides was developed. In this process, enolization of N-acyl iminosulfinamides was followed by O-silylation to generate O-silyl N-iminosulfinyl N,O-ketene aminal intermediates, which undergo a [2,3]-shift to afford α-sulfenylamino imidates that were converted to the corresponding carboxamides after desilylation triggered by acidic aqueous workup. Chirality is transferred from the sulfur stereocenter to the α-carbon, thereby enabling the enantioselective installation of an amino group at the α-position of amides.

Endoscopic transcortical expanded transforaminal transvenous transchoroidal approach to third ventricle lesion resection using an endoport.

OBJECTIVE: To investigate the clinical experience and application value of endoscopic resection of lesions in and around the third ventricle using a transcortical expanded transforaminal transvenous transchoroidal approach with an endoport. METHODS: Clinical data and follow-up results of seven patients who underwent the removal of lesions in the third ventricle and its adjacent area with an endoport-guided endoscopic system from January 2018 to December 2020 in the Department of Neurosurgery, Zhongshan Hospital Affiliated to Fudan University, were analyzed retrospectively. Two other patients from the Affiliated Pediatric Hospital of Fudan University and the Affiliated Hospital of Guizhou Medical University, respectively, were included in the analysis. RESULTS: A total of nine cases of third ventricle tumors were included in the study, including six women and three men, with an average age of 37.8 years (4-84 years old) and a follow-up time of 6-44 months. These nine tumor cases included two pilocytic astrocytomas, one diffuse midline glioma (H3 K27-altered), two craniopharyngiomas, two choroid plexus (CP) papillomas, one germinoma, and one pineal parenchymal tumor of intermediate differentiation. Total resection was completed in eight cases, with one near-total resection. There were no complications related to the surgical approach, such as epilepsy, aphasia, or hemiplegia. CONCLUSIONS: The endoscope transcortical expanded transforaminal transvenous transchoroidal approach using an endoport can safely and effectively remove third ventricle lesions. This approach can reach a wide area, from the anterior to the posterior third ventricle.

Chiral spiro phosphoric acid-catalysed enantioselective reaction of ketenes with N-H pyrroles.

In the presence of a chiral spiro phosphoric acid catalyst, the asymmetric reaction of disubstituted ketenes with N-H pyrroles occurred to afford enantioenriched C-acylated pyrroles bearing α-stereogenic carbon centres. The described reaction constitutes a rare example of a catalytic asymmetric reaction of ketenes with carbon-based nucleophiles.

Lentivirus-mediated RNAi silencing targeting ABCC2 increasing the sensitivity of a human nasopharyngeal carcinoma cell line against cisplatin.

BACKGROUND: High resistance to drug is taken as a characteristic of human tumors, which is usually mediated by multidrug resistance-associated genes. ABCC2, an ATP-binding cassette multidrug resistance transporter, is found to be expressed in a variety of human cancers. In this study the effect of a RNAi construct targeting ABCC2 on the chemosensitivity of NPC cell line CNE2 against cisplatin was investigated. METHODS: Lentiviral vectors were constructed to allow an efficient expression of anti-ABCC2 siRNA. The effective target sequence comprised nucleotides 1707-1727 of the human ABCC2 mRNA. The cell clones expressing the construct were picked and expanded, followed by identification using qRT-PCR and western blot method. As control, lentiviral vector containing invalid RNAi sequence was transfected to CNE2 cells. In vitro, cellular accumulation of cisplatin was detected by HPLC. The capacity of cellular growth and sensitivity of cells against cisplatin were detected by MTT assay. In vivo, the sensitivity of the tumor tissues against cisplatin were evaluated by transplanted CNE2 nude mice model. RESULTS: Two CNE2 cell clones with reduced expression of targeted ABCC2 mRNA and protein for more than 70% by qRT-PCR and western blot were established, and no differences were shown in proliferation rates compared to control CNE2 cells by growth curves analysis. In vitro the accumulation of intracellular cisplatin in these CNE2 cell clones with reduced expression of ABCC2 increased markedly, accompanied by increased sensitivity against cisplatin. In vivo, the growth of CNE2 solid tumors with a stably transfected anti-ABCC2 siRNA construct was significantly inhibited by cisplatin in transplanted nude mice model. CONCLUSION: Our investigation demonstrated that lentivirus-mediated RNAi silencing targeting ABCC2 might reverse the ABCC2-related drug resistance of NPC cell line CNE2 against cisplatin.

A regulatory mutant on TRIM26 conferring the risk of nasopharyngeal carcinoma by inducing low immune response.

The major histocompatibility complex (MHC) is most closely associated with nasopharyngeal carcinoma (NPC), but the complexity of its genome structure has proven challenging for the discovery of causal MHC loci or genes. We conducted a targeted MHC sequencing in 40 Cantonese NPC patients followed by a two-stage replication in 1065 NPC cases and 2137 controls of Southern Chinese descendent. Quantitative RT-PCR analysis (qRT-PCR) was used to detect gene expression status in 108 NPC and 43 noncancerous nasopharyngeal (NP) samples. Luciferase reporter assay and chromatin immunoprecipitation (ChIP) were used to assess the transcription factor binding site. We discovered that a novel SNP rs117565607_A at TRIM26 displayed the strongest association (OR = 1.909, Pcombined = 2.750 × 10-19 ). We also observed that TRIM26 was significantly downregulated in NPC tissue samples with genotype AA/AT than TT. Immunohistochemistry (IHC) test also found the TRIM26 protein expression in NPC tissue samples with the genotype AA/AT was lower than TT. According to computational prediction, rs117565607 locus was a binding site for the transcription factor Yin Yang 1 (YY1). We observed that the luciferase activity of YY1 which is binding to the A allele of rs117565607 was suppressed. ChIP data showed that YY1 was binding with T not A allele. Significance analysis of microarray suggested that TRIM26 downregulation was related to low immune response in NPC. We have identified a novel gene TRIM26 and a novel SNP rs117565607_A associated with NPC risk by regulating transcriptional process and established a new functional link between TRIM26 downregulation and low immune response in NPC.

RGC32 induces epithelial-mesenchymal transition by activating the Smad/Sip1 signaling pathway in CRC.

Response gene to complement 32 (RGC32) is a transcription factor that regulates the expression of multiple genes involved in cell growth, viability and tissue-specific differentiation. However, the role of RGC32 in tumorigenesis and tumor progression in colorectal cancer (CRC) has not been fully elucidated. Here, we showed that the expression of RGC32 was significantly up-regulated in human CRC tissues versus adjacent normal tissues. RGC32 expression was significantly correlated with invasive and aggressive characteristics of tumor cells, as well as poor survival of CRC patients. We also demonstrated that RGC32 overexpression promoted proliferation, migration and tumorigenic growth of human CRC cells in vitro and in vivo. Functionally, RGC32 facilitated epithelial-mesenchymal transition (EMT) in CRC via the Smad/Sip1 signaling pathway, as shown by decreasing E-cadherin expression and increasing vimentin expression. In conclusion, our findings suggested that overexpression of RGC32 facilitates EMT of CRC cells by activating Smad/Sip1 signaling.

A data-driven method for syndrome type identification and classification in traditional Chinese medicine.

The efficacy of traditional Chinese medicine (TCM) treatments for Western medicine (WM) diseases relies heavily on the proper classification of patients into TCM syndrome types. The authors developed a data-driven method for solving the classification problem, where syndrome types were identified and quantified based on statistical patterns detected in unlabeled symptom survey data. The new method is a generalization of latent class analysis (LCA), which has been widely applied in WM research to solve a similar problem, i.e., to identify subtypes of a patient population in the absence of a gold standard. A well-known weakness of LCA is that it makes an unrealistically strong independence assumption. The authors relaxed the assumption by first detecting symptom co-occurrence patterns from survey data and used those statistical patterns instead of the symptoms as features for LCA. This new method consists of six steps: data collection, symptom co-occurrence pattern discovery, statistical pattern interpretation, syndrome identification, syndrome type identification and syndrome type classification. A software package called Lantern has been developed to support the application of the method. The method was illustrated using a data set on vascular mild cognitive impairment.

Design and preparation of Epstein-Barr virus genome-wide cDNA probes.

OBJECTIVE: To design and clone all known and predicted coding genes of Epstein-Barr virus (EBV) as the cDNA probes for preparing the microarray for EBV detection, thereby to facilitate further investigation of the pathogenetic role of EBV. METHODS: Oligo 6.0 software, BLAST program and Primer Premier 5 software were employed to design and screen the cDNA probes of the whole EBV genome, whose length ranged from 300 to 600 mer each with high specificity. These cDNA probes obtained through PCR and reverse transcriptase (RT)-PCR amplification from the genomic DNA and RNA of B95-8 cells and nasopharyngeal carcinoma (NPC) tissue were cloned into T/A clone vector, followed by identification of these probes by sequencing analysis. RESULT AND CONCLUSION: A total of 85 gene fragments (BWRF1 gene-contained 7 repeats of open reading frames) coding for proteins and 2 EBERs in EBV genome were successfully cloned, not including LF1 and LF3 genes that did not exist in EBV genome of B95-8 cells, which provides the basis for preparing microarray to explore the role of EBV genome in its related diseases.

Effects of different concentrations of amino acids in the culture medium on preimplantation mouse embryo development in vitro.

OBJECTIVE: To evaluate the effects of amino acids (AA) on the development of in vitro cultured preimplantation embryos of Kunming mice, and define the optimal AA concentration for embryo culture. METHODS: Totally 630 zygotes were collected from the oviducts of superovulated female Kunming mice, which were cultured in protein-free potassium simplex optimized medium (mKSOM) supplemented with Eagle's essential amino acids and Eagle's non-essential amino acids of different concentrations (mKSOM, mKSOM+1/16AA, mKSOM+1/8AA, mKSOM+1/4AA, mKSOM+1/2AA, mKSOM+AA, and mKSOM+2AA). RESULTS: The embryos cultured with the amino acids showed higher development rate to both 8-cell embryo stage and blastocyst stage than those cultured without amino acids. The correlation of amino acid concentration with 8-cell and blastocyst development rates conformed to the cubic model, with the highest development rate to both of the two stages observed at half of the amino acid concentration. CONCLUSION: Amino acids can promote the development of preimplantation Kunming mouse embryos, but excessively high concentration of amino acids impair embryo development possibly because of metabolic and osmotic pressure changes of the embryos as well as toxicity of ammonium resulting from the metabolism of amino acids.