Black carbon and elemental characterization of PM2.5 in dense traffic areas in two cities in Fiji, a Small Island Developing State.

PM2.5 characterizations are essential in understanding its impact on the health of the exposed population. Sampled PM2.5 by Mani et al. (2020) was characterized to determine atmospheric metal concentration and inhalation health risk in Suva and Lautoka Cities, the only two cities in Fiji and one of the largest in the South Pacific Islands. Twenty-two elements (Al, As, Ba, Ca, Cd, Co, Cr, Cu, Fe, K, Mg, Mn, Mo, Na, Ni, P, Pb, S, Si, Sr, V, Zn) were analyzed using ICP-OES. Black Carbon (BC) sampling was also done at three different sites in Suva City, namely, Fiji National University Samabula Intersection site, Suva City Bus Station site and the Reservoir Road Community Settlement Site as well as at Lautoka City Bus Station. Mean BC concentrations over the sampling period were found to be 3.9 ± 2.9 (median = 3.3 µg/m3), 2.6 ± 2.7 µg/m3 (median = 1.7 µg/m3), 2.4 ± 2.3 µg/m3 (median = 1.7 µg/m3) and 4.0 ± 4.7 µg/m3 (median = 2.4 µg/m3) respectively. Health risk assessments (Carcinogenic Risk (CR) and Non-Carcinogenic Risk (HQ)) were also done to assess the risk of inhalation exposure in adults and children. The Hazard Index for children in Lautoka (HI = 1.03) was found to slightly exceed the safe level of 1. This study provides the first inventory of atmospheric particulate bound metal concentrations and diurnal BC profiles in Fiji and informs policy makers and scientists for further studies.

The Z-cad dual fluorescent sensor detects dynamic changes between the epithelial and mesenchymal cellular states.

BACKGROUND: The epithelial to mesenchymal transition (EMT) has been implicated in metastasis and therapy resistance of carcinomas and can endow cancer cells with cancer stem cell (CSC) properties. The ability to detect cancer cells that are undergoing or have completed EMT has typically relied on the expression of cell surface antigens that correlate with an EMT/CSC phenotype. Alternatively these cells may be permanently marked through Cre-mediated recombination or through immunostaining of fixed cells. The EMT process is dynamic, and these existing methods cannot reveal such changes within live cells. The development of fluorescent sensors that mirror the dynamic EMT state by following the expression of bona fide EMT regulators in live cells would provide a valuable new tool for characterizing EMT. In addition, these sensors will allow direct observation of cellular plasticity with respect to the epithelial/mesenchymal state to enable more effective studies of EMT in cancer and development. RESULTS: We generated a lentiviral-based, dual fluorescent reporter system, designated as the Z-cad dual sensor, comprising destabilized green fluorescent protein containing the ZEB1 3' UTR and red fluorescent protein driven by the E-cadherin (CDH1) promoter. Using this sensor, we robustly detected EMT and mesenchymal to epithelial transition (MET) in breast cancer cells by flow cytometry and fluorescence microscopy. Importantly, we observed dynamic changes in cellular populations undergoing MET. Additionally, we used the Z-cad sensor to identify and isolate minor subpopulations of cells displaying mesenchymal properties within a population comprising predominately epithelial-like cells. The Z-cad dual sensor identified cells with CSC-like properties more effectively than either the ZEB1 3' UTR or E-cadherin sensor alone. CONCLUSIONS: The Z-cad dual sensor effectively reports the activities of two factors critical in determining the epithelial/mesenchymal state of carcinoma cells. The ability of this stably integrating dual sensor system to detect dynamic fluctuations between these two states through live cell imaging offers a significant improvement over existing methods and helps facilitate the study of EMT/MET plasticity in response to different stimuli and in cancer pathogenesis. Finally, the versatile Z-cad sensor can be adapted to a variety of in vitro or in vivo systems to elucidate whether EMT/MET contributes to normal and disease phenotypes.

Phosphorylation of serine 367 of FOXC2 by p38 regulates ZEB1 and breast cancer metastasis, without impacting primary tumor growth.

Metastatic competence is contingent upon the aberrant activation of a latent embryonic program, known as the epithelial-mesenchymal transition (EMT), which bestows stem cell properties as well as migratory and invasive capabilities upon differentiated tumor cells. We recently identified the transcription factor FOXC2 as a downstream effector of multiple EMT programs, independent of the EMT-inducing stimulus, and as a key player linking EMT, stem cell traits and metastatic competence in breast cancer. As such, FOXC2 could serve as a potential therapeutic target to attenuate metastasis. However, as FOXC2 is a transcription factor, it is difficult to target by conventional means such as small-molecule inhibitors. Herein, we identify the serine/threonine-specific kinase p38 as a druggable upstream regulator of FOXC2 stability and function that elicits phosphorylation of FOXC2 at serine 367 (S367). Using an orthotopic syngeneic mouse tumor model, we make the striking observation that inhibition of p38-FOXC2 signaling selectively attenuates metastasis without impacting primary tumor growth. In this model, circulating tumor cell numbers are significantly reduced in mice treated with the p38 inhibitor SB203580, relative to vehicle-treated counterparts. Accordingly, genetic or pharmacological inhibition of p38 decreases FOXC2 protein levels, reverts the EMT phenotype and compromises stem cell attributes in vitro. We also identify the EMT-regulator ZEB1-known to directly repress E-cadherin/CDH1-as a downstream target of FOXC2, critically dependent on its activation by p38. Consistent with the notion that activation of the p38-FOXC2 signaling axis represents a critical juncture in the acquisition of metastatic competence, the phosphomimetic FOXC2(S367E) mutant is refractory to p38 inhibition both in vitro and in vivo, whereas the non-phosphorylatable FOXC2(S367A) mutant fails to elicit EMT and upregulate ZEB1. Collectively, our data demonstrate that FOXC2 regulates EMT, stem cell traits, ZEB1 expression and metastasis in a p38-dependent manner, and attest to the potential utility of p38 inhibitors as antimetastatic agents.

Inhibition of FOXC2 restores epithelial phenotype and drug sensitivity in prostate cancer cells with stem-cell properties.

Advanced prostate adenocarcinomas enriched in stem-cell features, as well as variant androgen receptor (AR)-negative neuroendocrine (NE)/small-cell prostate cancers are difficult to treat, and account for up to 30% of prostate cancer-related deaths every year. While existing therapies for prostate cancer such as androgen deprivation therapy (ADT), destroy the bulk of the AR-positive cells within the tumor, eradicating this population eventually leads to castration-resistance, owing to the continued survival of AR-/lo stem-like cells. In this study, we identified a critical nexus between p38MAPK signaling, and the transcription factor Forkhead Box Protein C2 (FOXC2) known to promote cancer stem-cells and metastasis. We demonstrate that prostate cancer cells that are insensitive to ADT, as well as high-grade/NE prostate tumors, are characterized by elevated FOXC2, and that targeting FOXC2 using a well-tolerated p38 inhibitor restores epithelial attributes and ADT-sensitivity, and reduces the shedding of circulating tumor cells in vivo with significant shrinkage in the tumor mass. This study thus specifies a tangible mechanism to target the AR-/lo population of prostate cancer cells with stem-cell properties.

Whom to blame for metastasis, the epithelial-mesenchymal transition or the tumor microenvironment?

Changes in the tumor microenvironment (TME) can trigger the activation of otherwise non-malignant cells to become highly aggressive and motile. This is evident during initial tumor growth when the poor vascularization in tumors generates hypoxic regions that trigger the latent embryonic program, epithelial-to-mesenchymal transition (EMT), in epithelial carcinoma cells (e-cars) leading to highly motile mesenchymal-like carcinoma cells (m-cars), which also acquire cancer stem cell properties. After that, specific bidirectional interactions take place between m-cars and the cellular components of TME at different stages of metastasis. These interactions include several vicious positive feedback loops in which m-cars trigger a phenotypic switch, causing normal stromal cells to become pro-tumorigenic, which then further promote the survival, motility, and proliferation of m-cars. Accordingly, there is not a single culprit accounting for metastasis. Instead both m-cars and the TME dynamically interact, evolve and promote metastasis. In this review, we discuss the current status of the known interactions between m-cars and the TME during different stages of metastasis and how these interactions promote the metastatic activity of highly malignant m-cars by promoting their invasive mesenchymal phenotype and CSC properties.

GD3 synthase regulates epithelial-mesenchymal transition and metastasis in breast cancer.

The epithelial-mesenchymal transition (EMT) bestows cancer cells with increased stem cell properties and metastatic potential. To date, multiple extracellular stimuli and transcription factors have been shown to regulate EMT. Many of them are not druggable and therefore it is necessary to identify targets, which can be inhibited using small molecules to prevent metastasis. Recently, we identified the ganglioside GD2 as a novel breast cancer stem cell marker. Moreover, we found that GD3 synthase (GD3S)--an enzyme involved in GD2 biosynthesis--is critical for GD2 production and could serve as a potential druggable target for inhibiting tumor initiation and metastasis. Indeed, there is a small molecule known as triptolide that has been shown to inhibit GD3S function. Accordingly, in this manuscript, we demonstrate that the inhibition of GD3S using small hairpin RNA or triptolide compromises the initiation and maintenance of EMT instigated by various signaling pathways, including Snail, Twist and transforming growth factor-ß1 as well as the mesenchymal characteristics of claudin-low breast cancer cell lines (SUM159 and MDA-MB-231). Moreover, GD3S is necessary for wound healing, migration, invasion and stem cell properties in vitro. Most importantly, inhibition of GD3S in vivo prevents metastasis in experimental as well as in spontaneous syngeneic wild-type mouse models. We also demonstrate that the transcription factor FOXC2, a central downstream effector of several EMT pathways, directly regulates GD3S expression by binding to its promoter. In clinical specimens, the expression of GD3S correlates with poor prognosis in triple-negative human breast tumors. Moreover, GD3S expression correlates with activation of the c-Met signaling pathway leading to increased stem cell properties and metastatic competence. Collectively, these findings suggest that the GD3S-c-Met axis could serve as an effective target for the treatment of metastatic breast cancers.

Malaysian undergraduates' knowledge and opinions on Early Childhood oral health.

AIM: In Malaysia, Early Childhood Caries (ECC) was found to affect 76.2% of 5-year-olds (2005). General practitioners are more likely to encounter preschool children and are in a better position to educate parents and caretakers about ECC and make appropriate referrals. This study assessed the knowledge and opinions on early childhood oral health among medical and dental undergraduates. MATERIALS AND METHODS: This cross sectional study involved 245 students enrolled in the first year medical (M1) and dental (D1) course and fourth year medical (M4) and dental (D4) course. The students completed a self-administered questionnaire which included knowledge and opinions on early childhood oral health. Comparisons between the groups were done using chi-square test. RESULTS: Dental students showed significantly better knowledge than medical students. D1 students showed significantly better knowledge of age of first tooth eruption over M1. Knowledge of recommended age for bottle weaning was higher among D4 students but not significantly more than M4 students. CONCLUSION: The majority of medical students showed inadequate knowledge indicating that medical curriculum should emphasise on oral health topics of public health relevance like ECC and its prevention. Dental students had better knowledge regarding early childhood oral health, but lacked knowledge on its preventive aspects.

Loss of breast epithelial marker hCLCA2 promotes epithelial-to-mesenchymal transition and indicates higher risk of metastasis.

Transition between epithelial and mesenchymal states is a feature of both normal development and tumor progression. We report that expression of chloride channel accessory protein hCLCA2 is a characteristic of epithelial differentiation in the immortalized MCF10A and HMLE models, while induction of epithelial-to-mesenchymal transition by cell dilution, TGFß or mesenchymal transcription factors sharply reduces hCLCA2 levels. Attenuation of hCLCA2 expression by lentiviral small hairpin RNA caused cell overgrowth and focus formation, enhanced migration and invasion, and increased mammosphere formation in methylcellulose. These changes were accompanied by downregulation of E-cadherin and upregulation of mesenchymal markers such as vimentin and fibronectin. Moreover, hCLCA2 expression is greatly downregulated in breast cancer cells with a mesenchymal or claudin-low profile. These observations suggest that loss of hCLCA2 may promote metastasis. We find that higher-than-median expression of hCLCA2 is associated with a one-third lower rate of metastasis over an 18-year period among breast cancer patients compared with lower-than-median (n=344, unfiltered for subtype). Thus, hCLCA2 is required for epithelial differentiation, and its loss during tumor progression contributes to metastasis. Overexpression of hCLCA2 has been reported to inhibit cell proliferation and is accompanied by increases in chloride current at the plasma membrane and reduced intracellular pH (pHi). We found that knockdown cells have sharply reduced chloride current and higher pHi, both characteristics of tumor cells. These results suggest a mechanism for the effects on differentiation. Loss of hCLCA2 may allow escape from pHi homeostatic mechanisms, permitting the higher intracellular and lower extracellular pH that are characteristic of aggressive tumor cells.

Knowledge, attitude and practice of oral health promoting factors among caretakers of children attending day-care centers in Kubang Kerian, Malaysia: a preliminary study.

BACKGROUND AND AIM: The role of caretakers at day-care centers has become more imperative in promoting oral health care in children since many new mothers opt to work outside their homes, leaving their children at day-care centers. The aim of this study is to assess the knowledge, attitude and practice of oral health promoting factors among secondary caretakers of children attending day-care centers. SETTINGS AND DESIGN: This was a cross-sectional exploratory study conducted among secondary caretakers in Kubang Kerian, Malaysia. MATERIALS AND METHODS: Thirty-four caretakers fulfilling the inclusion and exclusion criteria participated in the study. The data were collected using a self-administered questionnaire addressing various aspects of knowledge, attitude and practice of oral health in children. Analysis was done using SPSS version 12.0. RESULTS: The knowledge of factors causing dental caries was found to be good among majority of the caretakers, but the concepts of transmissibility of caries and effect of hidden sugars were not evident. Seventy one percent did not know that frequent bottle feeding could cause tooth decay. Attitudes seemed to be governed by the cultural practices of the region rather than the knowledge obtained. The knowledge was not translated to practice adequately. Giving sweetened liquid in bottles was practiced by 53% of the caretakers. CONCLUSION: Implementation of nursery-based oral health promotion programs for secondary caretakers is needed to counteract early childhood caries.

Oral health care in the elderly population in Malaysia--a review.

One of the undeniable facts about living is that everyday we are getting older. By 2050, it is projected that one out of every five Malaysians will be 60 or older. The economic challenge of this demographic change will affect development and the financial implications of sustaining well being of this group are formidable and complex. This population group has extensive oral disease, medical problems that complicate their oral care, and unique dental treatment challenges. The authors discuss the problem of oral healthcare among the aged, its impediments and propose some approaches for improvement to better serve the needs of this group of vulnerable members of our nation.

Phenobarbitone-mediated translocation of the cytosolic proteins interacting with the 5'-proximal region of rat liver CYP2B1/B2 gene into the nucleus.

The positive element (PE) (-69 to -98 bp) within the 5'-proximal region of the CYP2B1/B2 gene (+1 to -179 bp) of rat liver is essential for phenobarbitone (PB) response and gives a single major complex with the rat liver cytosol in gel shift analysis. This complex corresponds to complex I (top) of the three complexes given by the nuclear extracts. PB treatment of rats leads to a decrease in complex I formation with the cytosol and PE and an increase in the same with the nuclear extract in gel shift analysis. Both the changes are counteracted by simultaneous okadaic acid administration. The nuclear protein giving rise to complex I has been isolated and has an M(r) of 26 kDa. The cytosolic counterpart consists of two species, 26 and 28 kDa, as revealed by Southwestern blot analysis using labeled PE. It is concluded that PB treatment leads to the translocation accompanied by processing of the cytosolic protein species into the nucleus that requires protein dephosphorylation. It is suggested that PB may exert a global regulation on the transcription of many genes by modulating the phosphorylation status of different protein factors involved in transcriptional regulation.

Receptor-mediated gene delivery approach demonstrates the role of 5'-proximal DNA region in conferring phenobarbitone responsiveness to CYP2B2 gene in rat liver in vivo.

The phenobarbitone (PB) responsiveness of the 5'-proximal region of the CYP2B1/B2 gene was examined in detail with plasmid DNA constructs containing G-free cassette as reporter, using in vivo targeting of the same DNA constructs into rat liver as galactosylated-polylysine complexes. The contribution of the proximal region (-1 to -179 bp) and the positive element (-69 to -98 bp) identified earlier in this laboratory to PB responsiveness was assessed. The results obtained on PB treatment of rats subjected to receptor-mediated gene delivery to liver were conclusive and dramatic, with the control (saline-treated) rats manifesting very little expression of the reporter, reflecting the in vivo picture of CYP2B1/B2 gene expression. The positive element conferred PB responsiveness to homologous and heterologous promoters. Deletion of the positive element led to elimination of PB response. The entire -179 bp region was significantly more effective in responding to PB treatment than the region up to -98 bp, both containing one copy of the positive element. Thus, the positive element and its flanking sequences in the 5'-proximal region are involved in conferring PB responsiveness to the CYP2B1/B2 gene.

Sperm-mediated gene transfer into oocytes of the golden hamster: assessment of sperm function.

The possibility of sperm as a vehicle to deliver foreign DNA to oocytes was tested in hamsters. Epididymal spermatozoa, incubated with linearized plasmid DNA encoding ovine growth hormone (pCMXoGH), showed a spontaneous tendency to interact with DNA. Kinetics of sperm uptake of DNA was determined by using [32P]-labeled DNA. Spermatozoa took up the added DNA by 15-30 min and the uptake was inhibited by human seminal fluid in a dose dependent manner. Addition of DNA did not affect the functional competence of spermatozoa, in terms of their ability to undergo capacitation and acrosome reaction (34.5% +/- 2.2 vs 35% +/- 1.5). The fertilizing ability of DNA treated-spermatozoa from hamsters and humans was assessed by zona-free hamster egg penetration assay. Number of sperm penetrated per oocyte were 23 +/- 4.5 and 1.4 +/- 1.3 for hamster and human spermatozoa, respectively. Penetrated oocytes harbored sperm-treated DNA both with hamster (30.2 cpm/oocyte) and human (19.2 cpm/oocyte) spermatozoa. These results show that the hamster and human spermatozoa have a strong tendency to interact with exogenous (foreign) DNA and are able to transfer DNA to oocytes. Sperm may be used as a vector for DNA transfer and this approach has potential in the production of transgenic animals.

Calcium hydroxide as a root canal filling material in primary teeth--a pilot study.

A pilot study was carried out in five mandibular primary molars using calcium hydroxide (Ca(OH)2) Paste as root canal filling material to find out an alternative to the routinely used zinc oxide eugenol (ZnOE), which is non-resorbable and causes deflection of succedaneous teeth. The six month clinical and radiographic follow-up carried out at 2 months interval, revealed that the treated teeth with Ca(OH)2 as root canal filling material were successful, showing no pain and tenderness to percussion. A tendency for decrease in size of radiolucency was seen. Two teeth showed complete healing of the periradicular radiolucency. Depletion of Ca(OH)2 paste was seen from the root canals even prior to physiological resorption of roots in 2 out of 5 treated teeth.

Ultrasonic evaluation of portosystemic collateral circulation in portal hypertension.

The aim of the study was to evaluate portosystemic collateral circulation in relation to (1)individual etiological groups of portal hypertension., (2) Presence and size of esophageal varices, (3) esophageal sclerotherapy and (4) ascites. A prospective study of 101 patients of portal hypertension was carried out. Patients were divided into 4 etiological groups: Alcoholic cirrhosis (ALD) (38), Non-alcoholic cirrhosis (NALD) (35), non cirrhotic portal fibrosis (NCPF) (14) and extrahepatic portal vein obstruction (EHPVO) (14). Esophageal varices were assessed and graded endoscopically into 3 categories: no varix, small varices and large varices. Evaluation of portosystemic collateral circulation, other than esophageal varices was done ultrasonically. "Other" portosystemic collaterals (lienorenal, gastrorenal, dilated paraumbilical and umbilical veins, paraduodenal and gall bladdes varices) were seen in 26 out of 101 patients and more commonly in the non-cirrhotic groups (50%) [NCPF: 57.14%, EHPVO: 42.86%] than in the cirrhotic group (16.44%) [ALD: 13.5%, NALD: 20%]. Gall bladder varices were the only form of ectopic (extra esophagogastric) varices visualised with an overall incidence of 3.96%. Collateral shunts were seen more frequently in patients without varices (100%), than in patients with small varices (34.88%) or large varices (7.84%), and in patients having undergone esophageal sclerotherapy (57.14%). Collateral circulation did not contribute to the development of ascites. 37 patients with ascites did not have collateral shunts. We conclude portosystmic circulation plays a decompressive role in portal hypertension and prevents formation of esophageal varices or prevents them from increasing in size. It is seen more frequently in noncirrhotic patients and in those having undergone sclerotherapy and does not contribute to development of ascites.

Evaluation of splenomegaly in portal hypertension.

Splenomegaly is obvious in portal hypertension, but controversy still exists over the relationship between splenic size or size of esophageal varices. Previous methods to assess spleen size are less accurate than ultrasonic estimation of spleen size by splenic volumetric index (SVI). In a prospective study, we evaluated 101 consecutive patients with portal hypertension for spleen size measured ultrasonically by SVI, presence and size of esophageal varices, and etiology of portal hypertension. A total of 219 age-matched controls were evaluated ultrasonically to define a normal SVI. Splenomegaly defined by 1 or 2 standard deviations of normal SVI had high accuracy in predicting portal hypertension, presence of esophageal varices, and provided a clue to the etiology of portal hypertension. However, there was no correlation between spleen size and size of esophageal varices.

A model for the transcriptional regulation of the CYP2B1/B2 gene in rat liver.

The phenobarbitone-responsive minimal promoter has been shown to lie between nt -179 and nt + 1 in the 5' (upstream) region of the CYP2B1/B2 gene in rat liver, on the basis of the drug responsiveness of the sequence linked to human growth hormone gene as reporter and targeted to liver as an asialoglycoprotein-DNA complex in vivo. Competition analyses of the nuclear protein-DNA complexes formed in gel shift assays with the positive (nt -69 to -98) and negative (nt -126 to -160) cis elements (PE and NE, respectively) identified within this region earlier indicate that the same protein may be binding to both the elements. The protein species purified on PE and NE affinity columns appear to be identical based on SDS/PAGE analysis, where it migrates as a protein of 26-28 kDa. Traces of a high molecular weight protein (94-100 kDa) are also seen in the preparation obtained after one round of affinity chromatography. The purified protein stimulates transcription of a minigene construct containing the 179 nt on the 5' side of the CYP2B1/B2 gene linked to the I exon in a cell-free system from liver nuclei. The purified protein can give rise to all the three complexes (I, II, and III) with the PE, just as the crude nuclear extract, under appropriate conditions. Manipulations in vitro indicate that the NE has a significantly higher affinity for the dephosphorylated form than for the phosphorylated form of the protein. The PE binds both forms. Phenobarbitone treatment of the animal leads to a significant increase in the phosphorylation of the 26- to 28-kDa and 94-kDa proteins in nuclear labeling experiments followed by isolation on a PE affinity column. We propose that the protein binding predominantly to the NE in the dephosphorylated state characterizes the basal level of transcription of the CYP2B1/B2 gene. Phenobarbitone treatment leads to phosphorylation of the protein, shifting the equilibrium toward binding to the PE. This can promote interaction with an upstream enhancer through other proteins such as the 94-kDa protein and leads to a significant activation of transcription.

Fraunhofer diffraction patterns from uniformly illuminated square output apertures with noncentered square obscurations.

Theoretical Fraunhofer diffraction patterns are presented for uniformly illuminated square apertures with noncentered square obscurations. The energy within a given subtended solid angle in the far field is calculated. It is shown that the cornered-off-axis obscuration provides much more far-field energy in a given spot size than the centered obscuration for the same clear aperture area and total energy, for example, 82% more far-field energy in the first Airy square for 50% obscuration, thus providing superior performance for practical systems.