Occurrence of a Hybrid Between Taenia saginata and Taenia asiatica Tapeworms in Cambodia

Human infection with Taenia asiatica or a hybrid between Taenia saginata and T. asiatica has not been reported in Cambodia. We detected for the first time a hybrid form between T. saginata and T. asiatica in Preah Vihear Province, Cambodia. An adult tapeworm specimen, i.e., 75 cm long strobila without scolex, was expelled from a 27-year-old man after praziquantel medication and purging. It was morphologically indistinguishable between T. saginata and T. asiatica. Several proglottids were molecularly analyzed to confirm the tapeworm species. The mitochondrial gene encoding cytochrome c oxidase subunit 1 (cox1) and nuclear genes encoding elongation factor-1α (ef1) and ezrin-radixin-moesin (ERM)-like protein (elp) were sequenced, and a single-allele analysis was performed to confirm the haploid genotype. The results revealed that our sample showed a discrepancy between the mitochondrial and 2 nuclear genes. It possessed homozygous sequences typical of T. saginata at cox1 and ef1 loci. However, it was heterozygous at the elp locus, with 1 allele in T. asiatica (elpA) and 1 in T. saginata (elpC), which indicates that it is a hybrid between T. saginata and T. asiatica. The present results confirmed the presence of a hybrid between T. saginata and T. asiatica in Cambodia and strongly suggest the existence of also ‘pure’ T. asiatica in Cambodia.     

A Chrysin Derivative Suppresses Skin Cancer Growth by Inhibiting Cyclin-dependent Kinases

Chrysin (5,7-dihydroxyflavone), a natural flavonoid widely distributed in plants, reportedly has chemopreventive properties against various cancers. However, the anticancer activity of chrysin observed in in vivo studies has been disappointing. Here, we report that a chrysin derivative, referred to as compound 69407, more strongly inhibited EGF-induced neoplastic transformation of JB6 P⁺ cells compared with chrysin. It attenuated cell cycle progression of EGF-stimulated cells at the G₁ phase and inhibited the G₁/S transition. It caused loss of retinoblastoma phosphorylation at both Ser-795 and Ser-807/811, the preferred sites phosphorylated by Cdk4/6 and Cdk2, respectively. It also suppressed anchorage-dependent and -independent growth of A431 human epidermoid carcinoma cells. Compound 69407 reduced tumor growth in the A431 mouse xenograft model and retinoblastoma phosphorylation at Ser-795 and Ser-807/811. Immunoprecipitation kinase assay results showed that compound 69407 attenuated endogenous Cdk4 and Cdk2 kinase activities in EGF-stimulated JB6 P⁺ cells. Pulldown and in vitro kinase assay results indicated that compound 69407 directly binds with Cdk2 and Cdk4 in an ATP-independent manner and inhibited their kinase activities. A binding model between compound 69407 and a crystal structure of Cdk2 predicted that compound 69407 was located inside the Cdk2 allosteric binding site. The binding was further verified by a point mutation binding assay. Overall results indicated that compound 69407 is an ATP-noncompetitive cyclin-dependent kinase inhibitor with anti-tumor effects, which acts by binding inside the Cdk2 allosteric pocket. This study provides new insights for creating a general pharmacophore model to design and develop novel ATP-noncompetitive agents with chemopreventive or chemotherapeutic potency.     

Morphological and Molecular Diagnosis of Necator americanus and Ancylostoma ceylanicum Recovered from Villagers in Northern Cambodia

Human hookworm infections caused by adult Ancylostoma spp. and Necator americanus are one of the most important tropical diseases. We performed a survey of intestinal helminths using the Kato-Katz fecal examination technique targeting 1,156 villagers residing in 2 northern provinces (Preah Vihear and Stung Treng) of Cambodia in 2018. The results revealed a high overall egg positive rate of intestinal helminths (61.9%), and the egg positive rate of hookworms was 11.6%. Nine of the hookworm egg positive cases in Preah Vihear Province were treated with 5–10 mg/kg pyrantel pamoate followed by purging with magnesium salts, and a total of 65 adult hookworms were expelled in diarrheic stools. The adult hookworms were analyzed morphologically and molecularly to confirm the species. The morphologies of the buccal cavity and dorsal rays on the costa were observed with a light microscope, and the nucleotide sequences of mitochondrial cytochrome c oxidase subunit 1 (cox1) gene were analyzed. The majority of the hookworm adults (90.7%) were N. americanus, whereas the remaining 9.3% were Ancylostoma ceylanicum, a rare hookworm species infecting humans. The results revealed a high prevalence of hookworm infections among people in a northern part of Cambodia, suggesting the necessity of a sustained survey combined with control measures against hookworm infections.     

Cerebral Blood Volume Calculated by Dynamic Susceptibility Contrast-Enhanced Perfusion MR Imaging: Preliminary Correlation Study with Glioblastoma Genetic Profiles

Purpose

To evaluate the usefulness of dynamic susceptibility contrast (DSC) enhanced perfusion MR imaging in predicting major genetic alterations in glioblastomas.

Materials and Methods

Twenty-five patients (M:F = 13∶12, mean age: 52.1±15.2 years) with pathologically proven glioblastoma who underwent DSC MR imaging before surgery were included. On DSC MR imaging, the normalized relative tumor blood volume (nTBV) of the enhancing solid portion of each tumor was calculated by using dedicated software (Nordic TumorEX, NordicNeuroLab, Bergen, Norway) that enabled semi-automatic segmentation for each tumor. Five major glioblastoma genetic alterations (epidermal growth factor receptor (EGFR), phosphatase and tensin homologue (PTEN), Ki-67, O6-methylguanine-DNA methyltransferase (MGMT) and p53) were confirmed by immunohistochemistry and analyzed for correlation with the nTBV of each tumor. Statistical analysis was performed using the unpaired Student t test, ROC (receiver operating characteristic) curve analysis and Pearson correlation analysis.

Results

The nTBVs of the MGMT methylation-negative group (mean 9.5±7.5) were significantly higher than those of the MGMT methylation-positive group (mean 5.4±1.8) (p = .046). In the analysis of EGFR expression-positive group, the nTBVs of the subgroup with loss of PTEN gene expression (mean: 10.3±8.1) were also significantly higher than those of the subgroup without loss of PTEN gene expression (mean: 5.6±2.3) (p = .046). Ki-67 labeling index indicated significant positive correlation with the nTBV of the tumor (p = .01).

Conclusion

We found that glioblastomas with aggressive genetic alterations tended to have a high nTBV in the present study. Thus, we believe that DSC-enhanced perfusion MR imaging could be helpful in predicting genetic alterations that are crucial in predicting the prognosis of and selecting tailored treatment for glioblastoma patients.     

Inhibition of MAGEA2 regulates pluripotency,proliferation, apoptosis,and differentiation in mouse embryonic stem cells

Mouse embryonic stem cells (mESCs) exhibit self-renewal and pluripotency, can differentiate into all three germ layers, and serve as an essential model in stem cell research and for potential clinical application in regenerative medicine. Melanoma-associated antigen A2 (MAGEA2) is not expressed in normal somatic cells but rather in different types of cancer, especially in undifferentiated cells, such as in the testis, differentiating cells, and ESCs. However, the role of MAGEA2 in mESCs remains to be clarified. Accordingly, in this study, we examined the expression and functions of MAGEA2 in mESCs. MAGEA2 messenger RNA (mRNA) expression was decreased during mESCs differentiation. MAGEA2 function was then evaluated in knockdown mESC. MAGEA2 knockdown resulted in decreased pluripotency marker gene expression in mESCs consequent to increased Erk1/2 phosphorylation. Decreased MAGEA2 expression inhibited mESC proliferation via S phase cell cycle arrest with a subsequent decrease in cell cycle-associated genes Cdk1, Cdk2, Cyclin A1, Cyclin D1, and Cdc25a. Apoptotic mESCs markedly increased along with cleaved forms of caspases 3, 6, and 7 and PARP expression, confirming caspase-dependent apoptosis. MAGEA2 knockdown significantly decreased embryoid body size in vitro when cells were differentiated naturally and teratoma size in vivo, concomitant with decreased ectoderm marker gene expression. These findings suggested that MAGEA2 regulates ESC pluripotency, proliferation, cell cycle, apoptosis, and differentiation. The enhanced understanding of the regulatory mechanisms underlying diverse mESC characteristics will facilitate the clinical application of mESCs.     

Importance of region-specific epithelial rearrangements in mouse rugae development

Epithelial appendages on palatal rugae develop during mouse palatogenesis through epithelial thickening and pattern formation. Recently, the patterned formation of nine rugae was observed together with the specific expression patterns of Shh in rodents. However, no crucial evidence was found for a direct association between Shh expression and the distinct structural formation of rugae. In order to reveal possible relationships, we investigated the morphological changes of rugae and expression patterns of Shh directly by in vitro organ culture at embryonic day 13 (E13) for 2 days. To compare and examine the diverse growing aspects of the palate and rugae, we carefully observed the detailed morphogenesis, with cell proliferation of the rugae occurring between E13 and E14.5. After 2 days of cultivation at E13, DiI micro-injections revealed that the middle part of the palate, adjacent to the upper molar-forming region, contributed to the formation of the subsequent structure of rugae by extensive cell rearrangement and proliferation within the epithelium in the preferred anteroposterior direction. The results also defined the intimate relationship between Shh expression and rugae formation.     

The parthenogenetic activation of canine oocytes with Ca-EDTA by various culture periods and concentrations

In the present study, canine oocytes were exposed to various concentrations of and durations of exposure to EDTA saturated with Ca(2+) (Ca-EDTA), a cell membrane-impermeable metal ion chelator, to determine if parthenogenetic activation could be induced. When oocytes were cultured for 48 or 72 h in parthenogenetic activation medium (PAM) without Ca-EDTA (control) or PAM supplemented with 1 or 5mM Ca-EDTA, the highest rate of pronuclear formation (PN) was obtained in oocytes cultured in 1mM Ca-EDTA for 48 h (8.0%; P<0.05). There was no pronuclear formation in the control group (PAM without Ca-EDTA). Oocytes treated with 5mM Ca-EDTA for 48 h or 1mM Ca-EDTA for 72 h formed a parthenogenetic pronucleus (3.1 and 4.5, respectively). However, there was no pronuclear formation in oocytes treated with 5mM Ca-EDTA for 72 h. In summary, exposure to Ca-EDTA can induce pronuclear formation in canine oocytes.