Receptor Tyrosine Kinases: Principles and Functions in Glioma Invasion.

Protein tyrosine kinases are enzymes that are capable of adding a phosphate group to specific tyrosines on target proteins. A receptor tyrosine kinase (RTK) is a tyrosine kinase located at the cellular membrane and is activated by binding of a ligand via its extracellular domain. Protein phosphorylation by kinases is an important mechanism for communicating signals within a cell and regulating cellular activity; furthermore, this mechanism functions as an "on" or "off" switch in many cellular functions. Ninety unique tyrosine kinase genes, including 58 RTKs, were identified in the human genome; the products of these genes regulate cellular proliferation, survival, differentiation, function, and motility. Tyrosine kinases play a critical role in the development and progression of many types of cancer, in addition to their roles as key regulators of normal cellular processes. Recent studies have revealed that RTKs such as epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor (PDGFR), c-Met, Tie, Axl, discoidin domain receptor 1 (DDR1), and erythropoietin-producing human hepatocellular carcinoma (Eph) play a major role in glioma invasion. Herein, we summarize recent advances in understanding the role of RTKs in glioma pathobiology, especially the invasive phenotype, and present the perspective that RTKs are a potential target of glioma therapy.

Physical activity and incident dementia in older Japanese adults: The Okayama study.

OBJECTIVE: To evaluate the association between regular physical activity and the risk of incident dementia in older Japanese adults. METHODS: This was a retrospective cohort study performed in Okayama City, Japan. Overall, 51 477 older Japanese adults were followed from 2008 to 2014. A health checkup questionnaire was used to assess regular physical activity. The Dementia Scale of long-term care insurance was used as a measure of incident dementia. Cox proportional hazard models were used to calculate adjusted hazard ratios, with their 95% confidence intervals, for the incidence of dementia across the categories of physical activity. RESULTS: During a 7-year follow-up, 13 816 subjects were considered as having incident dementia. Compared with participants who performed physical activity less than or equal to one time per week, the multivariate adjusted hazard ratio values (95% confidence intervals) for participants who performed physical activity greater than or equal to two times per week but not every day and those who performed physical activity every day were 0.79 (0.75-0.84) and 0.94 (0.89-0.98), respectively. The interaction of physical activity and sex was statistically significant (P < .01). In subgroup analysis, the multivariate-adjusted hazard ratio values (95% confidence intervals) remained low, at 0.76 (0.70-0.84) in males and 0.81 (0.76-0.87) in females who performed physical activity greater than or equal to two times per week but not every day; they were 0.82 (0.76-0.89) in males and 1.01 (0.95-1.07) in females who performed physical activity every day. CONCLUSIONS: Regular physical activity could reduce the risk of incident dementia in older Japanese adults, except females who performed physical activity every day.

Strong therapeutic potential of γ-secretase inhibitor MRK003 for CD44-high and CD133-low glioblastoma initiating cells.

The Notch signal regulates both cell viability and apoptosis, and maintains stemness of various cancers including glioblastoma (GBM). Although Notch signal inhibition may be an effective strategy in treating GBM initiating cells (GICs), its applicability to the different subtypes of GBM remains unclear. Here, we analyzed the effectiveness of MRK003, a preclinical γ-secretase inhibitor, on GICs. Nine patient-derived GICs were treated by MRK003, and its efficacy on cell viability, apoptosis, sphere forming ability and Akt expression level which might be related to Notch downstream and be greatly important signals in GBM was evaluated. MRK003 suppressed viability and sphere-formation ability, and induced apoptosis in all GICs in varying doses of MRK003. Based on their sensitivities to MRK003, the nine GICs were divided into "relatively sensitive" and "relatively resistant" GICs. Sensitivity to MRK003 was associated with its inhibitory effect on Akt pathway. Transgenic expression of the myristoylated Akt vector in relatively sensitive GICs partially rescued the effect of MRK003, suggesting that the effect of MRK003 was, at least in part, mediated through inhibition of the Akt pathway. These GICs were differentiated by the expression of CD44 and CD133 with flow cytometric analysis. The relatively sensitive GICs are CD44-high and CD133-low. The IC50 of MRK003 in a set of GICs exhibited a negative correlation with CD44 and positive correlation with CD133. Collectively, MRK003 is partially mediated by the Akt pathway and has strong therapeutic potential for CD44-high and CD133-low GICs.

Fascin regulates chronic inflammation-related human colon carcinogenesis by inhibiting cell anoikis.

By a proteomics-based approach, we identified an overexpression of fascin in colon adenocarcinoma cells (FPCKpP-3) that developed from nontumorigenic human colonic adenoma cells (FPCK-1-1) and were converted to tumorigenic by foreign-body-induced chronic inflammation in nude mice. Fascin overexpression was also observed in the tumors arising from rat intestinal epithelial cells (IEC 6) converted to tumorigenic in chronic inflammation which was induced in the same manner. Upregulation of fascin expression in FPCK-1-1 cells by transfection with sense fascin cDNA converted the cells tumorigenic, whereas antisense fascin-cDNA-transfected FPCKpP-3 cells reduced fascin expression and lost their tumor-forming ability in vivo. The tumorigenic potential by fascin expression was consistent with their ability to survive and grow in the three-dimensional multicellular spheroids. We found that resistance to anoikis (apoptotic cell death as a consequence of insufficient cell-to-substrate interactions), which is represented by the three-dimensional growth of solid tumors in vivo, was regulated by fascin expression through caspase-dependent apoptotic signals. From these, we demonstrate that fascin is a potent suppressor to caspase-associated anoikis and accelerator of the conversion of colonic adenoma cells into adenocarcinoma cells by chronic inflammation.

Prognostic paradox: brain damage around the glioblastoma resection cavity.

Hyperintense lesions around the resection cavity on magnetic resonance diffusion-weighted imaging (MR-DWI) frequently appear after brain tumor surgery due to the damage of surrounding brain. The putative connection between the lesion and the prognosis for patients with glioblastoma (GBM) was explored. This retrospective study reviewed consecutive sixty-one patients with newly diagnosed GBM. Postoperative MRI was performed within 2 weeks after the initial surgery. We classified the cases into two groups depending on whether DWI hyperintense lesions were observed or not [DWI(+) group and DWI(-) group]. Progression-free survival (PFS) and overall survival (OS) were compared between the two groups. Forty-two patients were identified. The various extents of hyperintense lesions around the resection cavity were observed in 28/42 (66.7%) cases. In the DWI(+) and DWI(-) groups, median PFS was 10.0 [95% confidence interval (CI) 8.4-11.5] and 6.7 (95% CI 4.9-8.5) months, respectively (p = 0.042), and median OS was 18.0 (95% CI 12.2-23.8) and 17.0 (95% CI 15.7-18.3) months, respectively (p = 0.254). On multivariate analysis, the presence of DWI hyperintense lesion was more likely to be an independent predictor for 6-month PFS (p = 0.019; HR, 0.038; 95% CI 0.002-0.582). Tumor recurrence appeared outside the former DWI hyperintense lesion. Hyperintense lesions surrounding the resected GBM on MR-DWI might be a favorable prognostic factor in patients with GBM.

Detection of the development of late-onset idiopathic aqueductal stenosis (LIAS) by chronological magnetic resonance imaging: a case report.

Late-onset idiopathic aqueductal stenosis (LIAS) has been recognized as one of the clinical entities of adulthood hydrocephalus for decades, although there have been no radiological reports that show normal ventricular systems before the development of LIAS. We present here an adolescent case of LIAS with a previously normal ventricular system on magnetic resonance imaging (MRI). A 17-year-old boy had been suffering from chronic headaches and mild intellectual disability (MID) since he became a teenager, and this had prevented him from leading an ordinary school life. MRIs on admission showed triventriculomegaly from the aqueductal stenosis that had not been detected on previous MRIs, at least until the age of 6. An endoscopic third ventriculostomy was successfully performed, which improved both the headache and the MID. The developmental mechanism of LIAS remains unclear, although the membranous ependymal-like tissue observed in the aqueduct suggested the preceding existence of an inflammatory process around this region. To the best of our knowledge, this case was noteworthy because the development of LIAS was clearly demonstrated on MRI for the first time.

Reversible acute bilateral blindness resulting from a frontal brain tumor: a case report.

We experienced an unusual case of a 15-year-old girl who suffered acute bilateral blindness caused by a frontal lobe tumour. She underwent emergent operation, after which her vision recovered. This case emphasizes that a brain mass can cause sudden onset blindness and an emergency intervention might save the patient's sight.

Combined effect of interstitial-substitutional elements on dislocation dynamics in nitrogen-added austenitic stainless steels.

Combined addition of interstitial-substitutional elements has been acknowledged to contribute to the increase in the strengths of steels. For further improvements in mechanical properties, their atomic-scale interaction mechanisms with dislocations are required to be examined. In this study, both high-resolution transmission electron microscopy and atom-probe tomography were used to correlate interstitial-substitutional elements with dislocation characteristics in austenitic stainless steels. Three types of dislocation core structures are identified and associated with their strain fields as well as N and Cr atoms in the N-added steels. It is revealed that N atoms interact elastically with the dislocations, followed by the segregation of Cr atoms via the chemical interaction between N and Cr atoms. This insight significantly improves the understanding of the multiple alloying mechanism in metallic materials such as interstitial alloys and high-entropy alloys.

Transthyretin amyloid cardiomyopathy: Literature review and red-flag symptom clusters for each cardiology specialty.

Wild-type transthyretin amyloid cardiomyopathy (ATTRwt-CM) is a progressive and infiltrative cardiac disorder that may cause fatal consequences if left untreated. The estimated survival time from diagnosis is approximately 3-6 years. Because of the non-specificity of initial symptom manifestation and insufficient awareness among treating physicians, approximately one-third of patients with ATTRwt-CM are initially misdiagnosed with other cardiac diseases. Although heart failure (HF) is the most common initial manifestation of ATTRwt-CM, observed in nearly 70% of affected patients, patients may also present with other cardiologic symptoms, such as atrial fibrillation (AF) and aortic stenosis (AS). This non-specific and diverse nature of the initial ATTRwt-CM presentation indicates that various cardiology subspecialties are involved in patient diagnosis and management. Standard guideline-directed pharmacological treatment for HF is not recommended for patients with ATTRwt-CM because of its limited effectiveness. However, no established algorithms are available regarding HF management in this patient population. This literature review provides an overview of the red flags for ATTRwt-CM and research findings regarding HF management in this patient population. In addition to commonly recognized red flags for ATTRwt-CM (e.g., HF, AF and severe AS), published literature identified potential red flags such as coronary microvascular dysfunction. For HF management in patients with ATTRwt-CM, the use of mineralocorticoid receptor antagonists (MRAs) was reported as a well-tolerated option associated with a low discontinuation rate and reduced mortality. Although there is no concrete evidence for recommendations against sodium-glucose cotransporter 2 inhibitor (SGLT2i) administration, research supporting its use is limited to small-scale studies. Robust evidence is lacking for AF ablation, implantable cardioverter-defibrillators and cardiac resynchronization therapy. Based on the published findings and our clinical experience as Japanese ATTRwt-CM experts, red-flag symptom clusters for each cardiology specialty (HF, arrhythmia and ischaemia/structural heart disease) and a treatment scheme for HF management are presented. As this research area remains at an exploratory stage, our observations would require further discussion among experts worldwide.

Glycogen synthase kinase 3ß inhibition sensitizes human glioblastoma cells to temozolomide by affecting O6-methylguanine DNA methyltransferase promoter methylation via c-Myc signaling.

Glycogen synthase kinase 3ß (GSK3ß) is a serine/threonine protein kinase involved in human cancers including glioblastoma. We have previously demonstrated that GSK3ß inhibition enhances temozolomide effect in glioma cells. In this report, we investigated the molecular mechanisms of sensitization of glioblastoma cells to temozolomide by GSK3ß inhibition, focusing on O(6)-methylguanine DNA methyltransferase (MGMT) gene silencing. Glioblastoma tissues from patients treated with the GSK3ß-inhibiting drugs were subjected to immunohistochemistry and methylation-specific PCR assay. Human glioblastoma cell lines T98G, U138, U251 and U87 were treated with a small-molecule GSK3ß inhibitor, AR-A014418 or GSK3ß-specific small interfering RNA. The combined effect of temozolomide and AR-A014418 on cell proliferation was determined by AlamarBlue assay and an isobologram method. MGMT promoter methylation was estimated by methylation-specific PCR and MethyLight assay. MGMT gene expression was evaluated by real-time quantitative reverse transcriptase-PCR. c-Myc and DNA (cytosine-5)-methyltransferase 3A binding to the MGMT promoter was estimated by chromatin immunoprecipitation assay. GSK3ß inhibition decreased phosphorylation of glycogen synthase and reduced MGMT expression and increased MGMT promoter methylation in clinical tumors. In glioblastoma cell lines, GSK3ß inhibition decreased cell viability, enhanced temozolomide effect and downregulated MGMT expression with relevant changes in the methylation levels of the MGMT promoter. Here, we showed for the first time that c-Myc binds to the MGMT promoter with consequent recruitment of DNA (cytosine-5)-methyltransferase 3A, regulating the levels of MGMT promoter methylation. The results of this study suggest that GSK3ß inhibition enhances temozolomide effect by silencing MGMT expression via c-Myc-mediated promoter methylation.

Isolation and characterization of human breast cancer cells with SOX2 promoter activity.

Sex determining region Y-box 2 (SOX2) is well known as one of the "stemness" factors and is often expressed in cancers including breast cancer. In this study, we developed a reporter system using fluorescent protein driven by the promoter for SOX2 gene to detect and isolate living SOX2-positive cells. Using this system, we determined that SOX2 promoter activities were well correlated with SOX2 mRNA expression levels in 5 breast cancer cell lines, and that the cell population with positive SOX2 promoter activity (pSp-T(+)) isolated from one of the 5 cell lines, MCF-7 cells, showed a high SOX2 protein expression and high sphere-forming activity compared with very low promoter activity (pSp-T(low/-)). The pSp-T(+) population expressed higher mRNA levels of several stemness-related genes such as CD44, ABCB1, NANOG and TWIST1 than the pSp-T(low/-) population whereas the two populations expressed CD24 at similar levels. These results suggest that the cell population with SOX2 promoter activity contains cancer stem cell (CSC)-like cells which show different expression profiles from those of CSC-marker genes previously recognized in human breast cancers.

Receptor tyrosine kinases: principles and functions in glioma invasion.

Protein tyrosine kinases are enzymes that are capable of adding a phosphate group to specific tyrosines on target proteins. A receptor tyrosine kinase (RTK) is a tyrosine kinase located at the cellular membrane and is activated by binding of a ligand via its extracellular domain. Protein phosphorylation by kinases is an important mechanism for communicating signals within a cell and regulating cellular activity; furthermore, this mechanism functions as an "on" or "off" switch in many cellular functions. Ninety unique tyrosine kinase genes, including 58 RTKs, were identified in the human genome; the products of these genes regulate cellular proliferation, survival, differentiation, function, and motility. Tyrosine kinases play a critical role in the development and progression of many types of cancer, in addition to their roles as key regulators of normal cellular processes. Recent studies have revealed that RTKs such as epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor (PDGFR), c-Met, Tie, Axl, discoidin domain receptor 1 (DDR1), and erythropoietin-producing human hepatocellular carcinoma (Eph) play a major role in glioma invasion. Herein, we summarize recent advances in understanding the role of RTKs in glioma pathobiology, especially the invasive phenotype, and present the perspective that RTKs are a potential target of glioma therapy.

Transversal Survey of Emergency Medicine Policy and Quality Metrics in Japan's Regional Health Care Plans.

Introduction: It is essential to establish appropriate medical quality metrics and make improvements to safely and efficiently deliver optimum emergency medical services. The Ministry of Health, Labor and Welfare (MHLW) recommends prefectures to establish numerical quality metrics in their regional healthcare plans (RHCP). The 7th RHCP was issued by the MHLW in 2017 along with a notice of planning in covering the six-year period from 2018 to 2023. In this descriptive study, the emergency medicine policies in the 7th RHCP of each prefecture were analyzed from a quality improvement perspective. Method: The authors examined the chapters on emergency medicine in the RHCPs of 47 prefectural governments for the overall structure, cost-benefits, and connection to community-based integrated care systems. The type and number of clinical measures listed as numerical metrics and their classification methods were emphasized. Result: Regarding the overall plan structure, 40 prefectural governments began their description with an analysis of current surroundings. In total, 24 prefectural governments mentioned community-based integrated care systems but none mentioned cost-benefit analysis. Altogether, only 43 of 47 prefectural governments (91%) indicated numerical metrics. The maximum number of numerical targets for quality measures by prefecture was 19, the minimum was 0, and the median was 4 (IQR: 3-6.5); there were 220 metrics in total, with 82 structural, 96 process, and 42 outcome measures. Additionally, 13 prefectures (28%) classified quality measures according to the MHLW's guidance, 6 (13%) used their own classification manner, while the others did not classify their measures. Conclusions: There were significant differences in emergency medicine policies and quality metrics among the prefectural governments. Further research is needed to develop and establish more comprehensive and appropriate metrics based on a common methodology to improve the quality of emergency medicine.

Down-regulating Effect of a Standardized Extract of Cultured Lentinula edodes mycelia on Cortactin in Prostate Cancer Cells Is Dependent on Malignant Potential.

BACKGROUND/AIM: The incidence and mortality rates of prostate cancer have been increasing worldwide. Although prostate cancer cells grow slowly in the local original site, once the cancer cells spread to distant organs they grow rapidly and show very aggressive features. Cortactin is a protein that regulates the actin cytoskeleton and plays crucial roles in cancer metastasis. Up-regulated cortactin is correlated with the metastatic capacity of prostate cancer cells. AHCC®, a standardized extract of cultured Lentinula edodes mycelia, has been previously reported to have cortactin-down-regulating effects on human pancreatic cancer cells. In the present study, the effects of AHCC® treatment on cortactin levels in prostate cancer cells was evaluated. MATERIALS AND METHODS: LNCaP.FGC, DU145, and PC-3 are human prostate cancer cell lines. LNCaP.FGC is well differentiated, androgen-dependent, and poorly metastatic. DU145 is less differentiated, androgen-independent, and moderate metastatic. PC-3 is less differentiated, androgen-independent, and highly metastatic. The effects of AHCC® treatment on cortactin levels in prostate cancer cells was evaluated by western blot. RESULTS: In vitro AHCC® treatment decreased cortactin levels in LNCaP.FGC and DU145 cells but did not change those in PC-3 cells. CONCLUSION: AHCC® treatment down-regulated cortactin expression in poor and moderate metastatic LNCaP.FGC and DU145 cells but showed no effect on cortactin expression in the highly metastatic PC-3 cells. Further studies are required to elucidate the mechanism of the resistance to AHCC® treatment in highly metastatic PC-3 cells.

Prediction of high-grade meningioma by preoperative MRI assessment.

High-grade (World Health Organization grades II and III) meningiomas grow aggressively and recur frequently, resulting in a poor prognosis. Assessment of tumor malignancy before treatment initiation is important. We attempted to determine predictive factors for high-grade meningioma on magnetic resonance (MR) imaging before surgery. We reviewed 65 meningiomas (39 cases, benign; 26 cases, high-grade) and assessed four factors: (1) tumor-brain interface (TBI) on T1-weighted imaging (T1WI), (2) capsular enhancement (CapE), i.e., the layer of the tumor-brain interface on gadolinium-enhanced T1WI (T1Gd), (3) heterogeneity on T1Gd, and (4) tumoral margin on T1Gd. All four factors were useful in distinguishing high-grade from benign meningiomas, according to univariate analysis. On multivariate regression analysis, unclear TBI and heterogeneous enhancement were independent predictive factors for high-grade meningioma. In meningiomas with an unclear TBI and heterogeneous enhancement, the probability of high-grade meningioma was 98%. Our data suggest that this combination of factors obtained from conventional sequences on MR imaging may be useful to predict high-grade meningioma.

Association between carotid plaque composition assessed by multidetector computed tomography and cerebral embolism after carotid stenting.

INTRODUCTION: We aimed to assess the relationship between atherosclerotic carotid plaque composition analyzed using multidetector computed tomography (MDCT) and the appearance of new ischemic lesions detected by diffusion-weighted images (DWI) after carotid artery stenting (CAS). METHODS: We quantitatively and qualitatively analyzed plaque characteristics in carotid arteries using MDCT before CAS in 19 patients. Carotid plaques were expediently subdivided into four components with Hounsfield unit (HU) values of <0, 0-60, 60-130, and >600. The incidence of distal embolism was evaluated with DWI. Pearson's correlation analyses were used to assess the association between plaque composition and the incidence of cerebral embolization. RESULTS: Fifteen patients (79%) demonstrated new DWI lesions after CAS. High-signal DWIs were noted as follows: one in six patients, 2 ~ 5 in five patients, 6 ~ 10 in two patients, and >10 in two patients. The mean volumes of the plaque components for HU < 0, 0-60, 60-130, and >600 were 5.4, 200, 260, and 59 mm(3), respectively. There was a strong correlation between the number of high-signal DWI lesions in the ipsilateral side and the plaque volume of HU < 0 (r = 0.927; P < 0.0001). There was a moderate correlation between the number of high-signal DWI lesions and the plaque volume of HU 0-60 (r = 0.568; P = 0.0099) and the sum total of HU < 0 and HU 0-60 (r = 0.609; P = 0.0047). CONCLUSIONS: Quantitative and qualitative tissue characterization of carotid plaques using MDCT might be a useful predictor for silent ischemic lesions after CAS.

Dissecting aneurysms of the vertebral artery--angiographic patterns at the dissecting site on balloon test occlusion.

INTRODUCTION: At present, the risk of future hemorrhage or ischemic insult from vertebral artery (VA) dissection cannot be estimated from available imaging data. We investigated the relationship between symptoms and the angiographic patterns of the dissecting site on balloon test occlusion (BTO) to develop guidelines for clinical decision-making. METHODS: We retrospectively reviewed 18 patients with unilateral VA dissection who presented with subarachnoid hemorrhage (SAH) or cerebral infarction. We analyzed the angiographic findings at the dissecting site on contralateral VA angiograms during BTO of the affected VA, classified the angiographic patterns into two types, and compared the symptoms they presented. RESULTS: Patients with dissection opacified from the distal to the proximal side are more likely to present with cerebral infarction than SAH. Conversely, patients with dissection opacified from the proximal to the distal side had a significantly higher incidence of SAH. CONCLUSIONS: Angiographic findings at the dissecting site on contralateral VA angiograms during BTO of the affected VA are helpful for considering the treatment and prognosis of patients with VA dissecting aneurysms.

Identification of tumor-initiating cells in a highly aggressive brain tumor using promoter activity of nucleostemin.

Controversy remains over whether the cancer stem cell (CSC) theory applies to all tumors. To determine whether cells within a highly aggressive solid tumor are stochastically or hierarchically organized, we combined a reporter system where the nucleostemin (NS) promoter drives GFP expression (termed NS-GFP) with a mouse brain tumor model induced by retroviral Ras expression on a p16(Ink4a)/p19(Arf)-deficient background. The NS-GFP system allowed us to monitor the differentiation process of normal neural stem/precursor cells by analyzing GFP fluorescence intensity. In tumor-bearing mice, despite the very high frequency of tumorigenic cells, we successfully identified the NS-GFP(+) cells as tumor-initiating cells (T-ICs). The clonal studies conclusively established that phenotypical heterogeneity can exist among the cells comprising a genetically homogeneous tumor, suggesting that this aggressive brain tumor follows the CSC model. Detailed analyses of the NS-GFP(+) brain tumor cells revealed that T-ICs showed activation of the receptor tyrosine kinase c-Met, which functions in tumor invasiveness. Thus, the NS-GFP system provides a powerful tool to elucidate stem cell biology in normal and malignant tissues.

Does caregiver's social bonding enhance the health of their children?: the association between social capital and child behaviors.

Little is known about the association between social capital and child behaviors. This study aims to investigate that association. A complete population-based, cross-sectional survey was conducted for all the caregivers with preschool children in a rural town in Okayama prefecture in Japan. Two dimensions of individual-level social capital and unhealthy child behaviors were reported by parent-administered questionnaire. We analyzed 354 preschool children (57.6% of all children for whom questionnaires were completed). Children whose main caregiver had high cognitive social capital were 89% less likely to miss breakfast (odds ratio [OR]＝0.11;95% confidence interval [CI]:0.01-1.03). Children whose caregiver had high structural social capital were 71% less likely to wake up late (OR＝0.29;95% CI:0.12-0.71) and 78% less likely to skip tooth brushing more than once per day (OR＝0.22;95% CI:0.05-0.93). Both cognitive and structural social capital were negatively associated with unhealthy child behaviors. A further intervention study is needed to confirm the impact of social capital on child behavior.