Identification and verification of feature biomarkers associated with immune cells in recurrent pregnancy loss.

OBJECTIVE: The aim of this study was to investigate the causes, diagnostic markers, and treatment methods for recurrent pregnancy loss (RPL) using bioinformatics approaches. MATERIALS AND METHODS: Bioinformatics methods were utilized to analyze gene expression databases to identify key genes and modules associated with RPL. Weighted gene co-expression network analysis (WGCNA) was employed to identify gene sets related to maternal-fetal immunity. Gene set variation analysis (GSVA) and protein-protein interaction networks were used to explore signaling pathways and molecular interactions in RPL. Immune cell infiltration was assessed using single-sample gene set enrichment analysis (ssGSEA). RESULTS: Thirteen genes were identified as potential diagnostic markers, some of which were involved in placental amino acid transport, glucose absorption, and reactive oxygen species production. Several gene sets related to protein transport, steroid synthesis, and glycosaminoglycan degradation were found to be associated with RPL. Immune cell infiltration analysis found that CD56bright NK cells and monocytes showed significantly increased infiltration in RPL and were associated with key hub genes. The validation of hub genes, including PCSK5, CCND2, SLC5A3, RASAL1, MYZAP, MFAP4, and P2RY14, as potential diagnostic markers, showed promising value. CONCLUSIONS: This study contributes to a better understanding of the etiology of RPL and potential diagnostic markers. The identified immune-related gene sets, signaling pathways, and immune cell infiltrations provide valuable insights for future research and therapeutic advancements in RPL.

Paclitaxel-resistant related lncRNA DBH-AS1 promotes the proliferation and invasion of esophageal cancer.

OBJECTIVE: Chemoresistance is one of the main obstacles in the clinical treatment of cancer. However, secondary resistance to paclitaxel poses new challenges for cancer treatment. Long noncoding RNAs regulate cellular functions at different levels and mechanisms and play an important role in the biological behavior of tumors. MATERIALS AND METHODS: LncRNA microarrays were used to detect lncRNAs in Paclitaxel-resistant cells and corresponding parental cells. Cell counting kit 8 and Transwell analysis were used to test the effect of lncRNA on function. RESULTS: The expression of lncRNA DBH-AS1 in TE-4 TAX-R cells was significantly higher than that in TE-4 cells. Transwell analysis showed that the overexpression of lncRNA DBH-AS1 increased the invasion of Eca cells. Cell scratches and Transwell analysis showed that the overexpression of lncRNA DBH-AS1 in Eca cell culture supernatants promoted the migration and invasion of HUVEC. In addition, lncRNA DBH-AS1 relies on miR-21 to regulate the expression of YOD1. CONCLUSIONS: Paclitaxel-resistant lncRNA DBH-AS1 appears to promote ECa cell proliferation and invasion by acting as a ceRNA and regulating miR-21-5p /YOD1 signaling pathway.

Increased risk of temporomandibular joint disorders and craniofacial deformities in patients with juvenile idiopathic arthritis: a population-based cohort study.

Juvenile idiopathic arthritis (JIA) is an autoimmune disease that has been proposed to involve the temporomandibular joint (TMJ). The aim of this study was to identify the relationships between JIA, TMJ disorders, and craniofacial deformities. This cohort study included patients diagnosed with clinically active JIA between 1999 and 2013 through a nationwide longitudinal health registry. The primary outcome was the presence of a TMJ disorder. The secondary outcome was the presence of a JIA-associated craniofacial deformity. A total of 2791 patients with JIA were included in the case group; 11,164 propensity score-matched individuals without JIA were selected from the same database as controls. TMJ disorders were present in 142 individuals: 48 (1.72%) in the case group and 94 (0.84%) in the control group (relative risk 2.047, 95% confidence interval 1.446-2.898). Craniofacial deformities were present in 374 individuals: 112 (4.01%) in the case group and 262 (2.35%) in the control group (relative risk 1.722, 95% confidence interval 1.380-2.148). Patients with JIA showed a significantly greater likelihood of developing TMJ disorders and craniofacial deformities compared to matched controls.

Total knee replacement in osteoarthritis patients on reducing the risk of major adverse cardiac events: a 18-year retrospective cohort study.

OBJECTIVES: Osteoarthritis (OA) is a common degenerative joint disease, and total knee replacement (TKR) is a successful surgical intervention for knee OA treatment. However, the risks of mortality and major cardiovascular events (MACEs) in patients receiving TKR remain unclear. This study investigated the risks of mortality and MACEs in knee OA patients who received TKR. METHODS: For this population-based cohort study, the Longitudinal Health Insurance Database 2000 was used. Two million individuals with knee OA defined by ICD-9-CM codes who received physical therapy between 1999 and 2017 were selected. For propensity score matching (PSM), we considered the year of knee OA diagnosis, demographics, comorbidities, co-medications, and knee OA-related hyaluronic acid or physical therapy at baseline. After PSM, regression analyses were performed to assess the association of mortality or MACEs with TKR and non-TKR individuals. RESULTS: We identified patients (n = 189,708) with a new diagnosis of knee OA between 2000 and 2017. In total, 10,314 propensity-score-paired TKR and non-TKR individuals were selected. The PSM cohort algorithm revealed that the risk of mortality or MACEs was lower in the TKR group (adjusted hazard ratio: 0.791; 95% confidence interval: 0.755-0.830) than in the non-TKR group. CONCLUSIONS: Patients with knee OA who received TKR had decreased risks of mortality and MACEs than those who did not receive TKR. Moreover, the TKR group received a reduced dosage of nonsteroidal anti-inflammatory drugs at the 1-year follow-up.

The relationship between famine exposure in early life and left atrial enlargement in adulthood.

BACKGROUND: Increased left atrium diameter (LAD) is associated with an elevated risk of cardiovascular diseases. The relationship between nutrition status and left atrial enlargement (LAE) is still unclear. The present study aimed to investigate the association of famine exposure in early life with LAE in adulthood. METHODS: Participants were divided into non-exposed, fetal, early, middle and late childhood exposed groups according to birth data. LAE was defined when LAD was ≥3.9 cm in women and ≥4.1 cm in men, or ≥2.3 cm m-2 by a sex-independent cut-off normalised for body surface area. Multivariate logistic regression was performed to calculate the odds ratio (OR) and confidence interval (CI) between famine exposure and LAE. RESULTS: In total, 2522 [905 male, mean (SD) age 59.1 (3.65) years] subjects were enrolled, including 392 (15.5%) LAE subjects. The prevalence of LAE in non-exposed, fetal, early, middle and late childhood exposed groups was 55 (10.8%), 38 (11.2%), 88 (18.1%), 102 (16.7%) and 109 (19.0%), respectively. Compared to the non-exposed group, the ORs for LAE were in fetal (OR = 0.956, 95% CI = 0.605-1.500, P = 0.847), late (OR = 1.748, 95% CI = 1.208-2.555, P = 0.003), middle (OR = 1.647, 95% CI = 1.140-2.403, P = 0.008) and early (OR = 1.630, 95% CI = 1.116-2.399, P = 0.012) childhood exposed groups after adjusting potential cofounders. When stratified by gender, smoking, body mass index, hypertension and diabetes, we found that the effect of famine exposure on LAE was only modified by diabetes (Pinteraction  = 0.007). CONCLUSIONS: Famine exposure during childhood stage might increase the risk of LAE in adults, and this effect interacts with diabetes.

The role of mitochondrial DNA mutations in coronary heart disease.

Coronary heart disease (CHD) is a leading cause of death worldwide. It is a multifactorial disorder resulting from harmful interactions between genetic and environmental factors. Due to the central role of mitochondria in cellular energy homeostasis, there is growing evidence supporting the role of damage to mitochondrial components such as mitochondrial DNA (mtDNA) in the pathogenesis and progression of CHD. However, the molecular mechanisms linking mtDNA and CHD remains unknown. In terms of mutations, we found that mitochondrial transfer RNA (mt-tRNA) genes are hot spots for pathogenic mutations associated with CHD. These mutations cause structural and functional changes in tRNA; specifically, failure of tRNA metabolism may impair mitochondrial protein synthesis and lead to mitochondrial dysfunction responsible for CHD. This review provides a detailed summary of the mtDNA mutations that have been reported to be associated with CHD and further discusses the possible molecular mechanisms behind the involvement of these mtDNA mutations in CHD.

LncRNA AWPPH accelerates the progression of non-small cell lung cancer by sponging miRNA-204 to upregulate CDK6.

OBJECTIVE: This study aims to uncover the function of long non-coding RNA (lncRNA) AWPPH in the progression of non-small cell lung cancer (NSCLC) and the potential mechanism. PATIENTS AND METHODS: AWPPH and microRNA (miRNA-204) levels in NSCLC tissues and adjacent normal tissues were detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Kaplan-Meier curves were introduced for assessing overall survival in NSCLC patients expressing high or low level of AWPPH. Potential correlation between expression levels of AWPPH and miRNA-204 in NSCLC tissues was analyzed by Spearman correlation test. Through Dual-Luciferase reporter gene assay, the interaction among AWPPH, miRNA-204, and CDK6 was identified. Potential impacts of AWPPH/miRNA-204/CDK6 regulatory loop on mediating proliferative, migratory, and invasive capacities of A549 cells were evaluated through cell counting kit-8 (CCK-8) and transwell assay. RESULTS: Upregulated AWPPH and downregulated miRNA-204 were determined in NSCLC tissues. AWPPH level was negatively correlated to overall survival in NSCLC patients and miRNA-204 level in NSCLC tissues. Silence of AWPPH attenuated proliferative, migratory, and invasive capacities in A549 cells. MiRNA-204 was the downstream gene of AWPPH. Knockdown of miRNA-204 reversed the decreased viability, migratory, and invasive rates in A549 cells with AWPPH knockdown. In addition, CDK6 was the target gene of miRNA-204. Overexpression of miRNA-204 downregulated CDK6 level in A549 cells. The attenuated proliferative, migratory, and invasive capacities in A549 cells overexpressing miRNA-204 were reversed after CDK6 overexpression. CONCLUSIONS: LncRNA AWPPH serves as the miRNA-204 sponge to upregulate CDK6 level, thus aggravating the progression of NSCLC.

The Relationship between Telomere Length and Cancer Mortality: Data from the 1999-2002 National Healthy and Nutrition Examination Survey (NHANES).

OBJECTIVES: The association between telomeres length (TL) and cancer mortality is uncertain. We tested the hypotheses that long TL are associated with reduced cancer mortality. DESIGN: Prospective cohort study. SETTING: the National Health and Nutrition Survey (NHANES, 1999-2002). PARTICIPANTS: The analytic sample included adults (n = 7183) who had TL measurements. MEASUREMENTS: DNA was obtained via blood samples. Telomere length was assessed using the quantitative polymerase chain reaction method. RESULTS: During follow-up (0.08-12.7 person-years, median = 9.5 years), we observed 195 participants had cancer as causes of death. TL was negatively corelated with age, body mass index (BMI), systolic blood pressure (SBP), C-reactive protein (CRP), race, diabetes, hypertension, cardiovascular diseases (CVD) and cancer mortality, conversely, positively corelated with alcohol use, but not related to diastolic blood pressure (DBP) and smoking. Kaplan-Meier analysis revealed that TL was significantly associated with cancer mortality (log-rank, P <0.001). CONCLUSIONS: Our study expands upon previous evidence of a relationship between TL and cancer mortality. TL may be a useful tool for evaluating risk of cancer mortality in American adults.

Risk of systemic lupus erythematosus in patients with human papillomavirus infection: a population-based retrospective cohort study.

BACKGROUND: Viral infection contributing to systemic lupus erythematosus (SLE) development has been largely reported. However, the SLE risk in patients with human papillomavirus (HPV) infection is unknown. METHODS: Data were retrieved from the Longitudinal Health Insurance Database (2000) in Taiwan. We identified 43,567 patients with HPV infection and 174,268 age- and sex-matched uninfected controls from 2002 to 2012. Individuals were followed up from index date (first date of diagnosis with HPV) until the occurrence of SLE, at the end of the study (December 2013), or when they were withdrawn from the insurance program. The incidence rate ratio (IRR) was calculated using the univariate Poisson regression. The adjusted hazard ratios (aHRs) were calculated, and sensitive and subgroups analyses were also conducted. RESULTS: Compared with the non-HPV controls, the IRR of SLE in HPV patients was 1.52 (95% confidence interval (CI): 1.09-2.12). The risk of SLE in HPV-infected individuals was significantly high (aHR: 1.48, 95% CI: 1.06-2.06) after adjusting for age, sex, and comorbidities. Men aged between 16 and 45 years were more susceptible to developing SLE (aHR: 21.57, 95% CI: 2.52-184.60, p = 0.0051). CONCLUSION: Our study showed a significantly higher risk of SLE among HPV-infected patients, especially in men aged between 16 and 45 years.

Periodontitis and dental scaling associated with pyogenic liver abscess: A population-based case-control study.

BACKGROUND AND OBJECTIVES: The purpose of this study was to investigate the relationship between periodontitis, dental scaling (DS) and pyogenic liver abscesses (PLAs). MATERIAL AND METHODS: A nationwide population-based case-control study was applied using data from the National Health Insurance Research Database in Taiwan. We identified and enrolled 691 PLA patients, who were individually matched by age and sex to 2764 controls. RESULTS: Conditional logistic regression was applied to estimate adjusted odds ratios (aORs) in patients with exposure to periodontitis and DS before PLA. After adjusting for other confounding factors, periodontitis remained a risk factor for PLA among patients aged 20-40 years, with an aOR of 2.31 (95% confidence interval [CI] = 1.37-3.90, P = .0018). In addition, the average aOR for PLA was significantly lower among patients with one DS (aOR = 0.76, 95% CI = 0.59-0.96) and more than one DS (aOR = 0.61, 95% CI = 0.39-0.95) within 1 year before the index date. CONCLUSION: According to these results, we concluded that adult patients with periodontitis aged <50 years old are more at risk for PLA than controls, particularly when they have no DS. Moreover, from 20 years of age, non-periodontal patients subjected to at least 2 DS per year are less at risk for PLA than controls.

Mutation profile and associated clinical features in Chinese patients with cytogenetically normal acute myeloid leukemia.

INTRODUCTION: Cytogenetically normal acute myeloid leukemia (CN-AML), which accounted for nearly half of total AML patients, is a highly heterogeneous subset of AML. The specific genetic profile and the ethnic features of CN-AML are worth to be studied. METHODS: Using deep sequencing technology, we detected the mutation pattern of 39 genes in 152 Chinese CN-AML patients and analyzed their clinical features. RESULTS: A total of 503 mutations of 39 genes were identified in 145 (95.4%) patients, with the median number of 3 mutations per case. Nine genes (NPM1, CEBPA, DNMT3A, GATA2, NRAS, TET2, FLT3, IDH2, and WT1) mutated in more than 10% patients. Function groups of myeloid transcription factors, activated signaling, and DNA methylation were most affected. The distribution of variant allele frequencies (VAF) of recurrent genes was different among functional groups. High mutation rates of CEBPA and GATA2 together with the low frequency of FLT3-ITD mutation seemed to be the distinct characteristics of Chinese patients. Furthermore, CEBPAbi and GATA2 were found to mutate most in M2 subtype, while NPM1 and DNMT3A mutated more in M4 and M5. The prognostic analysis identified CEBPAmo mutation as an inferior factor. FLT3-ITD, TP53, DNMT3A, CEBPAmo, and WT1 mutations were selected as high-risk markers to identify the CN-AML patients with poor prognosis. CONCLUSION: Our study provided the valuable information of ethnic genetic characteristics and the clinical relevance of Chinese CN-AML patients.

Capacitive deionization of a RO brackish water by AC/graphene composite electrodes.

A feasibility study for water recycling and reuse of a reverse osmosis (RO) brackish wastewater by capacitive deionization (CDI) was carried out in the present work. Palm-shell wastes enriched in carbon was recycled to yield valuable activated carbon (AC) that has advantages of high surface area, high specific capacitance, and low electrical resistance as the CDI electrodes. The GAC prepared by dispersion of AC in the graphene (rGO) layers has a high surface area and electrical conductivity for CDI. The GAC electrodes have increasing electrosorption efficiencies from 1.6 to 3.0% during the repeated electrosorption-regeneration cycles under +1.2 â†’ 0 → +1.2 V while the efficiencies the AC electrodes decrease from 2.7 to 1.6%. It is clear that the GAC-based electrodes have a better electrosorption efficiency and stability in, for example, the three repeated electrosoption-regeneration cycles for CDI of the wastewater. This work also exemplifies that the AC recycled from biomass such as palm-shell wastes can be used in CDI electrodes for recycling and reuse of wastewater.

Analysis and control of the photon beam position at PLS-II.

At third-generation light sources, the photon beam position stability is a critical issue for user experiments. In general, photon beam position monitors are developed to detect the real photon beam position, and the position is controlled by a feedback system in order to maintain the reference photon beam position. At Pohang Light Source II, a photon beam position stability of less than 1 µm r.m.s. was achieved for a user service period in the beamline, where the photon beam position monitor is installed. Nevertheless, a detailed analysis of the photon beam position data was necessary in order to ensure the performance of the photon beam position monitor, since it can suffer from various unknown types of noise, such as background contamination due to upstream or downstream dipole radiation, and undulator gap dependence. This paper reports the results of a start-to-end study of the photon beam position stability and a singular value decomposition analysis to confirm the reliability of the photon beam position data.

A transverse bunch by bunch feedback system for Pohang Light Source upgrade.

The Pohang Light Source upgrade (PLS-II) project has successfully upgraded the Pohang Light Source (PLS). The main goals of the PLS-II project are to increase the beam energy to 3 GeV, increase the number of insertion devices by a factor of two (20 IDs), increase the beam current to 400 mA, and at the same time reduce the beam emittance to below 10 nm by using the existing PLS tunnel and injection system. Among 20 insertion devices, 10 narrow gap in-vacuum undulators are in operation now and two more in-vacuum undulators are to be installed later. Since these narrow gap in-vacuum undulators are most likely to produce coupled bunch instability by the resistive wall impedance and limit the stored beam current, a bunch by bunch feedback system is implemented to suppress coupled bunch instability in the PLS-II. This paper describes the scheme and performance of the PLS-II bunch by bunch feedback system.

Induced maturation of hepatic progenitor cells in vitro

Hepatic progenitor cells (HPCs) are a potential cell source for liver cell transplantation but do not function like mature liver cells. We sought an effective and reliable method to induce HPC maturation. An immortalized HP14.5 albumin promoter-driven Gaussian luciferase (ALB-GLuc) cell line was established from HPCs isolated from fetal mouse liver of post coitus day 14.5 mice to investigate the effect of induction factors on ALB promoter. HP14.5 parental cells were cultured in DMEM with different combinations of 2% horse serum (HS), 0.1 µM dexamethasone (DEX), 10 ng/mL hepatic growth factor (HGF), and/or 20 ng/mL fibroblast growth factor 4 (FGF4). Trypan blue and crystal violet staining were used to assess cell proliferation with different induction conditions. Expression of hepatic markers was measured by semi-quantitative RT-PCR, Western blot, and immunofluorescence. Glycogen storage and metabolism were detected by periodic acid-Schiff and indocyanine green (ICG) staining. GLuc activity indicated ALB expression. The combination of 2% HS+0.1 µM Dex+10 ng/mL HGF+20 ng/mL FGF4 induced the highest ALB-GLuc activity. Cell proliferation decreased in 2% HS but increased by adding FGF4. Upon induction, and consistent with hepatocyte development, DLK, AFP, and CK19 expression decreased, while ALB, CK18, and UGT1A expression increased. The maturity markers tyrosine aminotransferase and apolipoprotein B were detected at days 3 and 6 post-induction, respectively. ICG uptake and glycogen synthesis were detectable at day 6 and increased over time. Therefore, we demonstrated that HPCs were induced to differentiate into functional mature hepatocytes in vitro, suggesting that factor-treated HPCs may be further explored as a means of liver cell transplantation.

Evaluation of adhesion force and binding affinity of phytohemagglutinin erythroagglutinating to EGF receptor on human lung cancer cells.

PHA-E is a natural product extracted from red kidney beans, and it has been reported to induce cell apoptosis by blocking EGFR in lung cancer cells. Because EGF is the major in vivo competitor to PHA-E in clinical application, PHA-E must be proved that has better affinity to EGFR than EGF. This study would focus on how PHA-E tightly bind to EGFR and the results would compare with EGF. The adhesion force, measured by AFM, between EGFR and PHA-E was 207.14±74.42 pN that was higher than EGF (183.65±86.93 pN). The equilibrium dissociation constant of PHA-E and EGF to EGFR was 2.4 10(-9)±1.4 10(-9) and 7.3 10(-8)±2.7 10(-8), respectively, that could evaluate binding affinity. The result showed that binding affinity of PHA-E to EGFR was one order higher than EGF to EGFR. In the results of flow cytometer and confocal microscope, we found binding efficiency of EGF to EGFR was decrease as the concentration of PHA-E increased. In the analysis of Western blot, treatment of A-549 cells with PHA-E resulted in a dose-dependent decrease in EGFR phosphorylation. In conclusion, we found that PHA-E had better adhesion force and binding affinity to EGFR than that of the EGF. The interaction between PHA-E and EGFR could block EGF binding and then inhibit EGFR phosphorylation. PHA-E could be developed into a new target molecule for lung cancer treatment that could be immobilized on the drug carrier to guide therapeutic particles to the tumor site.

Characterization of the major histocompatibility complex class I A alleles in cynomolgus macaques of Vietnamese origin.

Cynomolgus macaques (Macaca fascicularis, Mafa) have emerged as an important animal model for infectious disease and transplantation research. Extensive characterization of their major histocompatibility complex (MHC) polymorphism regions therefore becomes urgently required. In this study, we identified 41 MHC class I A nucleotide sequences in 34 unrelated cynomolgus macaques of Vietnamese origin farmed in Southern China, including eight novel Mafa-A sequences. We found two sequences with perfect identity and six sequences with close similarity to previously defined MHC class I alleles from other populations, especially from Indonesian-origin macaques. We also found three Vietnamese-origin cynomolgus macaque MHC class I sequences for which the predicted protein sequences identical throughout their B and F binding pockets to Mamu-A1*001:01 and Mamu-A3*13:03, respectively. This is important because Mamu-A1*001:01 and Mamu-A3*13:03 are associated with longer survival and lower set-point viral load in simian immunodeficiency virus (SIV)-infected rhesus monkeys. These findings have implications for the evolutionary history of Vietnamese-origin cynomolgus macaque as well as for the use of this model in SIV/SHIV (a virus combining parts of the HIV and SIV genomes) research.

SIRT3 regulates mitochondrial protein acetylation and intermediary metabolism.

The sirtuins are a family of nicotinamide adenine dinucleotide (NAD(+))-dependent protein deacetylases that regulate cell survival, metabolism, and longevity. Humans have seven sirtuins (SIRT1-SIRT7) with distinct subcellular locations and functions. SIRT3 is localized to the mitochondrial matrix and its expression is selectively activated during fasting and calorie restriction. Activated SIRT3 deacetylates several key metabolic enzymes-acetyl-coenzyme A synthetase, long-chain acyl-coenzyme A (acyl-CoA) dehydrogenase (LCAD), and 3-hydroxy-3-methylglutaryl CoA synthase 2-and enhances their enzymatic activity. Disruption of SIRT3 activity in mice, either by genetic ablation or during high-fat feeding, is associated with accelerated development of metabolic abnormalities similar to the metabolic syndrome in humans. SIRT3 is therefore emerging as a metabolic sensor that responds to change in the energy status of the cell and modulates the activity of key metabolic enzymes via protein deacetylation.

MAP kinase dependent cyclinE/cdk2 activity promotes DNA replication in early sea urchin embryos.

Sea urchins provide an excellent model for studying cell cycle control mechanisms governing DNA replication in vivo. Fertilization and cell cycle progression are tightly coordinated by Ca(2+) signals, but the mechanisms underlying the onset of DNA replication after fertilization remain less clear. In this study we demonstrate that calcium-dependent activation of ERK1 promotes accumulation of cyclinE/cdk2 into the male and female pronucleus and entry into first S-phase. We show that cdk2 activity rises quickly after fertilization to a maximum at 4 min, corresponding in timing to the early ERK1 activity peak. Abolishing MAP kinase activity after fertilization with MEK inhibitor, U0126, substantially reduces the early peak of cdk2 activity and prevents cyclinE and cdk2 accumulation in both sperm pronucleus and zygote nucleus in vivo. Both p27(kip1) and roscovitine, cdk2 inhibitors, prevented DNA replication suggesting cdk2 involvement in this process in sea urchin. Inhibition of cdk2 activity using p27(kip1) had no effect on the phosphorylation of MBP by ERK, but completely abolished phosphorylation of retinoblastoma protein, a cdk2 substrate, indicating that cdk2 activity is downstream of ERK1 activation. This pattern of regulation of DNA synthesis conforms to the pattern observed in mammalian somatic cells.