Biological and prognostic significance of NDRG2 downregulation in oral squamous cell carcinoma.

OBJECTIVE: Downregulation of N-myc downstream-regulated gene 2 (NDRG2), a tumor suppressor gene, has been associated with poor clinical outcomes in various cancers. However, the prognostic significance of NDRG2 in oral squamous cell carcinoma (OSCC) remains unknown. This study aimed to evaluate the prognostic value of NDRG2 downregulation in OSCC and to elucidate the mechanism by which NDRG2 is downregulated and the biological role of NDRG2 in tumor progression. METHODS: Immunohistochemical and in silico analyses of NDRG2 expression were performed, and the correlation between NDRG2 expression and clinicopathological data was analyzed. The effect of NDRG2 knockdown on the biological behavior of OSCC cells was investigated and the effect of 5-aza-2'-deoxycytidine (5-aza-dC) on NDRG2 expression was determined. RESULTS: NDRG2 expression was significantly downregulated and DNA hypermethylation of NDRG2 was frequently found in head and neck SCC, including OSCC. Low NDRG2 expression was significantly correlated with adverse clinicopathological features and worse survival in OSCC. NDRG2 knockdown could enhance the oncogenic properties of OSCC cells. NDRG2 mRNA levels in OSCC cells could be restored by 5-aza-dC. CONCLUSION: Downregulation of NDRG2 promotes tumor progression and predicts poor prognosis in OSCC. Therefore, restoration of NDRG2 expression may be a potential therapeutic strategy in OSCC.

Association between Genetic Polymorphism of GSTP1 and Toxicities in Patients Receiving Platinum-Based Chemotherapy: A Systematic Review and Meta-Analysis.

Platinum-based chemotherapy regimens have been proven to be effective in various cancers; however, considerable toxicities may develop and can even lead to treatment discontinuation. Diverse factors may influence adverse treatment events, with pharmacogenetic variations being one prime example. Polymorphisms within the glutathione S-transferase pi 1 (GSTP1) gene may especially alter enzyme activity and, consequently, various toxicities in patients receiving platinum-based chemotherapy. Due to a lack of consistency in the degree of elevated complication risk, we performed a systematic literature review and meta-analysis to determine the level of platinum-associated toxicity in patients with the GSTP1 rs1695 polymorphism. We conducted a systematic search for eligible studies published before January 2022 from PubMed, Web of Science, and EMBASE based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated to evaluate the strength of the association between the rs1695 polymorphism and various toxicities. Ten eligible studies met the inclusion criteria. The pooled ORs for hematological toxicity and neutropenia in the patients with the variant (G) allele were 1.7- and 2.6-times higher than those with the AA genotype (95% CI 1.06-2.73 and 1.07-6.35), respectively. In contrast, the rs1695 polymorphism resulted in a 44% reduced gastrointestinal toxicity compared to wild-type homozygotes. Our study found that the GSTP1 rs1695 polymorphism was significantly correlated with platinum-induced toxicities. The study also revealed that rs1695 expression exhibited tissue-specific patterns and thus yielded opposite effects in different tissues. A personalized chemotherapy treatment based on these polymorphisms may be considered for cancer patients in the future.

Elevated Risk of Fluoropyrimidine-Associated Toxicity in European Patients with DPYD Genetic Polymorphism: A Systematic Review and Meta-Analysis.

Background: Fluoropyrimidine is widely used owing to its clinical efficacy, however, patients with dihydropyrimidine dehydrogenase (DPD) deficiency can experience fluoropyrimidine-associated toxicity. The dihydropyrimidine dehydrogenase (DPYD) gene encodes DPD, and studies suggest that DPYD polymorphisms can result in DPD deficiency. Since there is not a complete consistency of how much the risk of complication is elevated, we aimed to conduct a systematic literature review and a meta-analysis to provide the risk of fluoropyrimidine-associated toxicity in patients with DPYD rs1801160 polymorphism. Methods: We searched for qualifying studies published before October 2021 from PubMed, Web of Science, and EMBASE based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated to evaluate the strength of the association between rs1801160 polymorphism and toxicities. A sensitivity analysis using the leave-one-out method was performed on the overall toxicity. Results: The pooled OR for overall toxicity in the patients with A allele was elevated 1.73 times higher than those with the GG genotype (95% CI 1.44-2.07). Sensitivity analysis yielded similar results, showing the robustness of the result. Subjects with variants showed a 2.37-fold increased hematological toxicity (95% CI 1.48-3.81); especially a 1.87-fold increased neutropenia compared to patients with wildtype (95% CI 1.49-2.34). Patients with A allele revealed 1.22 times higher gastrointestinal toxicity compared to those with GG genotype (95% CI 0.93-1.61), and among gastrointestinal toxicity, the risk of diarrhea was elevated 1.43 times higher in those with variants than patients with wildtype (95% CI 1.12-1.83). Conclusions: rs1801160 polymorphism is associated with elevated fluoropyrimidine-associated toxicity. Therefore, rs1801160 can be a potential candidate for DPD deficiency screening prior to fluoropyrimidine-based regimen.

Factors Associated with Thyroid-Related Adverse Events in Patients Receiving PD-1 or PD-L1 Inhibitors Using Machine Learning Models.

Targets of immune checkpoint inhibitors (ICIs) regulate immune homeostasis and prevent autoimmunity by downregulating immune responses and by inhibiting T cell activation. Although ICIs are widely used in immunotherapy because of their good clinical efficacy, they can also induce autoimmune-related adverse events. Thyroid-related adverse events are frequently associated with anti-programmed cell death 1 (PD-1) or anti-programmed cell death-ligand 1 (PD-L1) agents. The present study aims to investigate the factors associated with thyroid dysfunction in patients receiving PD-1 or PD-L1 inhibitors and to develop various machine learning approaches to predict complications. A total of 187 patients were enrolled in this study. Logistic regression analysis was conducted to investigate the association between such factors and adverse events. Various machine learning methods were used to predict thyroid-related complications. After adjusting for covariates, we found that smoking history and hypertension increase the risk of thyroid dysfunction by approximately 3.7 and 4.1 times, respectively (95% confidence intervals (CIs) 1.338-10.496 and 1.478-11.332, p = 0.012 and 0.007). In contrast, patients taking opioids showed an approximately 4.0-fold lower risk of thyroid-related complications than those not taking them (95% CI 1.464-11.111, p = 0.007). Among the machine learning models, random forest showed the best prediction, with an area under the receiver operating characteristic of 0.770 (95% CI 0.648-0.883) and an area under the precision-recall of 0.510 (95%CI 0.357-0.666). Hence, this study utilized various machine learning models for prediction and showed that factors such as smoking history, hypertension, and opioids are associated with thyroid-related adverse events in cancer patients receiving PD-1/PD-L1 inhibitors.

Association between the Arylalkylamine N-Acetyltransferase (AANAT) Gene and Seasonality in Patients with Bipolar Disorder.

OBJECTIVE: Bipolar disorder (BD) is complex genetic disorder. Therefore, approaches using clinical phenotypes such as biological rhythm disruption could be an alternative. In this study, we explored the relationship between melatonin pathway genes with circadian and seasonal rhythms of BD. METHODS: We recruited clinically stable patients with BD (n=324). We measured the seasonal variation of mood and behavior (seasonality), and circadian preference, on a lifetime basis. We analyzed 34 variants in four genes (MTNR1a, MTNR1b, AANAT, ASMT) involved in the melatonin pathway. RESULTS: Four variants were nominally associated with seasonality and circadian preference. After multiple test corrections, the rs116879618 in AANAT remained significantly associated with seasonality (corrected p=0.0151). When analyzing additional variants of AANAT through imputation, the rs117849139, rs77121614 and rs28936679 (corrected p=0.0086, 0.0154, and 0.0092) also showed a significant association with seasonality. CONCLUSION: This is the first study reporting the relationship between variants of AANAT and seasonality in patients with BD. Since AANAT controls the level of melatonin production in accordance with light and darkness, this study suggests that melatonin may be involved in the pathogenesis of BD, which frequently shows a seasonality of behaviors and symptom manifestations.

Psychopathologic structure of bipolar disorders: exploring dimensional phenotypes, their relationships, and their associations with bipolar I and II disorders.

BACKGROUND: Given its diverse disease courses and symptom presentations, multiple phenotype dimensions with different biological underpinnings are expected with bipolar disorders (BPs). In this study, we aimed to identify lifetime BP psychopathology dimensions. We also explored the differing associations with bipolar I (BP-I) and bipolar II (BP-II) disorders. METHODS: We included a total of 307 subjects with BPs in the analysis. For the factor analysis, we chose six variables related to clinical courses, 29 indicators covering lifetime symptoms of mood episodes, and 6 specific comorbid conditions. To determine the relationships among the identified phenotypic dimensions and their effects on differentiating BP subtypes, we applied structural equation modeling. RESULTS: We selected a six-factor solution through scree plot, Velicer's minimum average partial test, and face validity evaluations; the six factors were cyclicity, depression, atypical vegetative symptoms, elation, psychotic/irritable mania, and comorbidity. In the path analysis, five factors excluding atypical vegetative symptoms were associated with one another. Cyclicity, depression, and comorbidity had positive associations, and they correlated negatively with psychotic/irritable mania; elation showed positive correlations with cyclicity and psychotic/irritable mania. Depression, cyclicity, and comorbidity were stronger in BP-II than in BP-I, and they contributed significantly to the distinction between the two disorders. CONCLUSIONS: We identified six phenotype dimensions; in addition to symptom features of manic and depressive episodes, various comorbidities and high cyclicity constructed separate dimensions. Except for atypical vegetative symptoms, all factors showed a complex interdependency and played roles in discriminating BP-II from BP-I.

Changes in Dietary Quality among Vietnamese Women Immigrants in Korea and Comparison with Korean Women.

The dietary behavior of immigrants starts changing upon their arrival in a new country. We evaluated changes in dietary quality of Vietnamese women immigrants in Korea and compared dietary quality with that of Korean women. Fifty-six Vietnamese women immigrants and 56 age-matched Korean women were recruited. Dietary quality were assessed using index of nutritional quality (INQ) and diet quality index-international (DQI-I). Dietary habits were assessed according to 4 dietary behaviors: a prudent, calorie control, dietary fat control, and sodium or salt control diet. DQI-I scores of Vietnamese immigrants decreased after immigration, especially the moderation score, although the variety score increased. Scores were significantly lower than those of Korean subjects (45.1 vs. 64.5; p < 0.001). Vietnamese women immigrants had significantly poorer nutrient balance and calorie intake control, although their fat and sodium control was better than that of Korean woman (p < 0.001). INQs of protein, niacin, phosphorus, iron, zinc were lower in immigrants who had lived longer in Korea than more recent immigrants (p < 0.05). Lower INQs of protein, fiber, vitamin A, B1, B6, C, folate, and phosphorus were related to higher body fat in Vietnamese immigrants (p < 0.05). In conclusion, dietary quality of Vietnamese immigrants decreased after migration, and dietary intake was inadequate compared with that of Korean women. In addition, diet quality of Vietnamese immigrants decreased with length of residence in Korea. There was a negative correlation between diet quality and body fat percent in Vietnamese women immigrants. Findings from this study may help improve diet quality and prevent obesity in Vietnam women immigrants.

Effects of genetic variants of ST8SIA2 and NCAM1 genes on seasonal mood changes and circadian preference in the general population.

ST8SIA2 and NCAM1 are functionally related genes forming polysialic acid (PSA) - neural cell adhesion molecule (NCAM) complex in suprachiasmatic nucleus (SCN), the regulating site of circadian biological rhythm. In this study, the relationship of ST8SIA2 and NCAM1 with circadian and seasonal rhythms of human behavior was explored. Subjects were 261 healthy Korean adults who were free of any history of clinically significant psychiatric symptoms. The phenotypes were circadian preference and seasonal change of mood and behavior (seasonality) measured by the Composite Scale of Morningness and the Seasonal Pattern Assessment Questionnaire, respectively. Thirty-four single nucleotide polymorphisms (SNPs) across the ST8SIA2 region and 15 SNPs of NCAM1 were analyzed. A nominally significant association with seasonality and circadian preference was observed in 21 variants of both genes. After corrections for multiple testing, associations of 8 SNPs of ST8SIA2 and 2 SNPs of NCAM1 with seasonality remained significant. Some of these SNPs were also associated with psychiatric disorders in previous studies. This study demonstrated a meaningful and/or suggestive evidence of association between behavioral phenotypes reflecting human biological rhythm and two interplaying genes involved in the plasticity of SCN's neuronal network.

Long-term response to mood stabilizer treatment and its clinical correlates in patients with bipolar disorders: a retrospective observational study.

BACKGROUND: The efficacy and utility of long-term prophylactic treatment in patients with bipolar disorders (BDs) have not been fully explored. This study aims to estimate the long-term clinical response of patients with BDs to mood stabilizer treatment and to identify the clinical factors associated with that response. METHODS: The study subjects consisted of 80 patients with bipolar I or bipolar II disorder who had been receiving treatment with lithium and/or valproate for more than 2 years at a single bipolar disorder clinic. The long-term response to the best treatment option based on treatment algorithms was evaluated using the Alda scale. Clinical characteristics were evaluated on a lifetime basis. Patients were classified into two response groups based on frequentist mixture analysis using the total Alda scale score. RESULTS: Thirty-four percent of the patients were good responders, with a total Alda score of 5 or higher. The treatment response rate did not differ between the lithium and valproate groups, but lithium and valproate combination therapy was associated with poorer response. The number of previous mixed episodes was associated with a worse response (p = 0.026). Of individual symptoms, delusions during manic episodes (p = 0.008) and increased appetite (p = 0.035) during depressive episodes were more common in moderate/poor responders than in good responders. Co-morbid anxiety disorders were more frequently observed in the moderate/poor response group (p = 0.008). CONCLUSIONS: Psychotic, mixed, and atypical features of BDs were found to be correlated with long-term treatment outcomes. Lithium and valproate showed similar efficacy but moderate/poor responders preferred to use polypharmacy.

Protein Interacting with Never in Mitosis A-1 Induces Glutamatergic and GABAergic Neuronal Differentiation in Human Dental Pulp Stem Cells.

INTRODUCTION: The purpose of this study was to investigate the role of protein interacting with never in mitosis A-1 (PIN1) in the neuronal or glial differentiation of human dental pulp stem cells (hDPSCs) and whether PIN1 can regulate determination of neuronal sub-phenotype. METHODS: After magnetic-activated cell sorting to separate CD34(+)/c-kit(+)/STRO-1(+) hDPSCs, cells were cultured in neurogenic medium. Differentiation was measured as Nissl staining and marker protein or mRNA expression by reverse transcriptase polymerase chain reaction, immunofluorescence, and flow cytometric analysis. RESULTS: PIN1 mRNA levels were upregulated in a time-dependent fashion during neurogenic differentiation. The PIN1 inhibitor juglone suppressed neuronal differentiation but promoted glial differentiation as assessed by the number of Nissl-positive cells and mRNA expression of neuronal markers (nestin, ßIII-tubulin, and NeuN) and a glial marker (glial fibrillary acidic protein). Conversely, overexpression of PIN1 by infection with adenovirus-PIN1 increased neuronal differentiation but decreased glial differentiation. Moreover, PIN1 overexpression increased the percentage of glutamatergic and GABAergic cells but decreased that of dopaminergic cells among total NeuN-positive hDPSCs. CONCLUSIONS: This is the first study to demonstrate that PIN1 overexpression induced glutamatergic and GABAergic neuronal differentiation but suppressed glial differentiation of hDPSCs, suggesting that enhancing PIN expression is important to obtain human glutamatergic and GABAergic neurons from hDPSCs.

In Vivo Inhibition of Porphyromonas gingivalis Growth and Prevention of Periodontitis With Quorum-Sensing Inhibitors.

BACKGROUND: Autoinducer (AI)-2 has an important role in biofilm formation in the oral environment. Mature biofilms formed as a result of the cell-to-cell communication make it difficult to overcome periodontitis with the use of antibiotics. Previous in vitro studies suggest that quorum-sensing inhibitors (QSIs) interfere with AI-2. This study compares the QSI effects resulting from an oral inoculation of Porphyromonas gingivalis in an experimental animal model. METHODS: Forty-five male mice were divided into three groups (n = 15 each): 1) infection; 2) QSI; and 3) control. Infection and QSI groups received oral inoculation of P. gingivalis, whereas treatment with QSIs (furane compound and d-ribose) was only performed in the QSIs group. The control group was a negative control not receiving manipulation. After 42 days, mice were sacrificed, and the distance from the alveolar bone crest (ABC) to the cemento-enamel junction (CEJ) was measured by microcomputed tomography. P. gingivalis DNA was quantified in the soft and hard tissues around the molar teeth by real-time polymerase chain reaction. RESULTS: Distance from ABC to CEJ was significantly increased in the P. gingivalis infection group compared with the control group (P = 0.02) and significantly decreased in the QSI group compared with the infection group (P = 0.02). The QSI group contained 31.64% of the bacterial DNA count of the infection group. CONCLUSION: Use of QSIs in the mice infection model showed a reduction of bone breakdown and a decrease in the number of bacteria in vivo, suggesting that QSIs can be a new approach to prevention and treatment of periodontitis.

KiSS-1 expression in oral squamous cell carcinoma and its prognostic significance.

Downregulated expression of KiSS-1 has been correlated with tumor progression, metastasis, and patient prognosis in various human malignancies. However, there is no information regarding the expression of KiSS-1 in oral squamous cell carcinoma (OSCC). Our aims were to examine KiSS-1 expression in OSCC tissue samples and cell lines and to determine its prognostic significance. KiSS-1 expression was significantly lower in lymph node (LN) metastases than in primary tumor tissues. Five of six OSCC cell lines showed absence or relatively low expression of KiSS-1. Correlations between KiSS-1 expression and clinicopathological parameters were statistically assessed. There were significant correlations between KiSS-1 expression and LN metastasis (p = 0.007), TNM stage (p = 0.024), and local recurrence (p = 0.012). In the Kaplan-Meier survival analysis, negative KiSS-1 expression significantly correlated with poorer overall survival (OS) and disease-free survival (DFS) (p = 0.000 and 0.000, respectively). Multivariate analysis using Cox regression modeling revealed that KiSS-1 expression was an independent prognostic factor for both OS and DFS (p = 0.001 and 0.000, respectively). Our findings suggested that KiSS-1 downregulation may play a role in tumor progression and metastasis of OSCC and may be a reliable biomarker for predicting clinical outcome in OSCC.

Complication After Extraction of Natal Teeth with Continued Growth of a Dental Papilla.

The purposes of this case report were to describe a growing two-cm gingival mass that developed after natal teeth were extracted in a four-month-old female patient, present a review of the literature on the growth of a gingival mass after the extraction of natal teeth, and illustrate the clinical and histological features that differentiate this condition from other types of gingival masses in infants. Histological examination of the excised mass revealed that it contained tooth-like hard tissue (regular and irregular dentin) that intermingled with bone, dental pulp, and fibrous tissue. We found eight cases from 1962 to 2009 in which a soft-tissue mass with dentin-like hard tissue or a tooth-like structure had developed after the extraction of natal teeth. Based on clinical and histological findings, we deduced that the mass was the result of abnormal growth of a residual dental papilla, including mesenchymal stem cells. Consequently, dentists, obstetricians, gynecologists, and pediatricians should be aware of this potential complication and observe caution before they extract natal teeth.

Durable Urushiol-Based Nanofilm with Water Repellency for Clear Overlay Appliances in Dentistry.

With increased esthetic needs, orthodontics is an indispensable medical treatment in dentistry, and transparent clear overlay appliances (COAs) are in general use to fix teeth. However, COAs are easily worn out because of the lack of durability. Here, we applied a nanofilm onto COAs using urushiol (U), a durable coating material from plant via a layer-by-layer assembly technique. In particular, polymerized urushiol (PU) provided COAs with higher mechanical strength in the large-scale assessment, lower cytotoxicity, and intrinsic hydrophobicity for antimicrobial use. In this report, we inceptively attempted to functionalize COAs with nanofilm for advanced biomedical use.

Non-infectious in-cell HIV-1 protease assay utilizing translocalization of a fluorescent reporter protein and apoptosis induction.

This study describes a non-infectious in-cell imaging assay for HIV-1 protease inhibitor screening. It is based on re-distribution of a fluorescence reporter protein upon protease cleavage and the fact that HIV-infected cells undergo apoptosis. The in-cell assay utilizes fluorescent reporter proteins consisting of an intracellular translocation signal sequence, a caspase-3-specific cleavage sequence, and a fluorescent tagging protein. The reporter proteins are designed to change their intracellular localization upon cleavage, either from the cytosol to a subcellular organelle (type I) or from a subcellular organelle to the cytosol (type II). Inhibition of protease activity can be monitored at the single cell level. Interestingly, the expression of HIV-1 protease induced endogenous caspase-3 activation; thus, the fluorescence reporter protein containing the caspase-3 cleavage sequence translocalized upon cleavage. This is the first time that HIV-1 protease expression, not whole virus infection of the cell, was observed to trigger the apoptotic pathway, including caspse-3 activation. A validation of this assay was performed with a known HIV-1 protease inhibitor, Ac-Leu-Val-phenylalanine. The clear cellular change in fluorescence pattern makes this system an ideal tool for various types of life science and drug discovery research, including high throughput and high content screening applications.

Cholangiographic characteristics of common bile duct dilatation in children.

AIM: To investigate whether children with congenital common bile duct dilatation (CBDD) differ from children with obstructive CBDD in cholangiographic characteristics. METHODS: In this retrospective cohort study, the baseline data and the results of imaging analyses were reviewed among children who had endoscopic retrograde cholangiopancreatography (ERCP) due to CBDD. ERCP was performed on all pediatric patients by experienced pediatric endoscopists. The maximal transverse diameter of the common bile duct (CBD) was measured on ERCP. To assess whether age-adjusted CBDD could be used for differential diagnosis, a CBDD severity index (SI) was calculated by dividing the measured CBD diameter by the age-corrected maximal diameter of a normal CBD. RESULTS: A retrospective medical chart review revealed that 85 consecutive children under 16 years of age with hepatobiliary disease and CBDD were referred to Seoul Asan Medical Center. Fifty-five (64.7%) children had congenital CBDD and 30 (35.3%) had obstructive CBDD. The two groups did not differ significantly in terms of clinical characteristics except for sex. The congenital and obstructive CBDD groups did not differ significantly in terms of mean CBD diameter (19.3 ± 9.6 mm vs 12.2 ± 4.1 mm, P > 0.05). However, congenital CBDD cases had a significantly higher mean SI than obstructive CBDD cases (3.62 ± 1.64 vs 1.98 ± 0.71, P = 0.01). In multivariate analysis, an SI value ≥ 2.32 and comorbidity with anomalous union of pancreaticobiliary duct (APBDU) in ERCP independently predicted congenital CBDD. CONCLUSION: Measuring the CBD may aid the differential diagnosis of both CBDD and APBDU in children.

Effects of genetic variations in NRG1 on cognitive domains in patients with schizophrenia and healthy individuals.

BACKGROUND: The neuregulin 1 (NRG1) gene has been investigated as a candidate susceptibility gene for schizophrenia. A number of studies have also explored the genetic effect of NRG1 on cognitive deficits related to schizophrenia, and thus far generated inconsistent results. The current study aimed to determine whether genetic variations in NRG1 are associated with cognitive domains in schizophrenic patients and healthy individuals. MATERIALS AND METHODS: Comprehensive neuropsychological tests composed of six cognitive domains were administered to 135 clinically stable patients with schizophrenia and 119 healthy individuals. On the basis of previous reports of positive association, a total of four single nucleotide polymorphisms were analyzed. In testing the genotype effect on cognitive domains, we used repeated-measure analysis for six cognitive domain scores of each individual as repeated measurements. RESULTS: An association of P-value less than 0.05 with at least one cognitive domain in patients and/or healthy individuals was observed for all of the single nucleotide polymorphisms. After applying the correction for multiple testing, the association remained statistically significant between rs6994992 and general cognitive ability (g) in the patient group and between rs2439272 and the 'working memory' domain in the group of healthy participants. CONCLUSION: This study suggests the involvement of NRG1 in the susceptibility for developing cognitive deficits in schizophrenic patients. For some cognitive domains, its genetic effect was also significant in generating interindividual variability within the normal functional range.

CD33-Targeted Lipid Nanoparticles (aCD33LNs) for Therapeutic Delivery of GTI-2040 to Acute Myelogenous Leukemia.

CD33-targeted lipid nanoparticles (aCD33LNs) were synthesized for delivery of GTI-2040, an antisense oligonucleotide (ASO) against the R2 subunit of ribonucleotide reductase, to acute myelogenous leukemia (AML). These LNs incorporated a deoxycholate-polyethylenimine (DOC-PEI) conjugate, which has shown significant activity to facilitate oligonucleotide delivery. Anti-CD33 scFv (aCD33) was added as a targeting ligand. The delivery efficiency of this system was investigated both in vitro and in vivo. When cells were treated with aCD33LN/GTI-2040, significant uptake was observed in CD33 positive Kasumi-1 cells. aCD33LNs loaded with GTI-2040 induced significant down-regulation of R2 mRNA and protein levels in AML cells. Moreover, aCD33LN/GTI-2040 showed a 15-fold reduction in the IC50 of antileukemic drug Ara-C in Kasumi-1 cells. In Kasumi-1 xenograft model, aCD33LN/GTI-2040 showed significant R2 downregulation compared to LN/GTI-2040. Furthermore, aCD33LN/GTI-2040 coadministered with Ara-C was shown to be highly effective in tumor growth inhibition and to greatly increase survival time of mice bearing Kasumi-1 xenograft tumors. The conjugate DOC-PEI has shown an ability to include calcein release from lipid nanoparticles, suggesting a potential mechanism contributing to efficient endosome release by DOC-PEI2K. These results indicate that aCD33LNs are a highly effective vehicle for the therapeutic delivery of antisense agents to AML.

Comparison of the effects of human dental pulp stem cells and human bone marrow-derived mesenchymal stem cells on ischemic human astrocytes in vitro.

This study assesses the cytoprotective effects of human dental pulp stem cells (hDPSCs) and conditioned medium from hDPSCs (CM-hDPSCs) on ischemic human astrocytes (hAs) in vitro compared with human bone marrow-derived mesenchymal stem cells (hMSCs). Ischemia of hAs was induced by oxygen-glucose deprivation (OGD). CM-hDPSCs and hMSCs were collected after 48 hr of culture. Cell death was determined by 3-[4,5-dimethylthialzol-2-yl]-2,5-diphenyltetrazolium bromide and cellular ATP assays. The expression of glial fibrillary acidic protein (GFAP) and musashi-1 as markers of reactive astrogliosis was examined with immunochemical staining. mRNA expression and reactive oxygen species (ROS) were analyzed by RT-PCR and flow cytometry, respectively. OGD increased cytotoxicity in a time-dependent manner and decreased cellular ATP content concomitantly in hAs. Pretreatment and posttreatment with hDPSCs were associated with greater recovery from OGD-induced cytotoxicity in hAs compared with hMSCs. Similarly, CM-hDPSCs had a greater effect on OGD-induced cytotoxicity in a dose-dependent manner. Pre- and posttreatment with CM-hDPSCs or CM-hMSCs attenuated OGD-induced GFAP, nestin, and musashi-1 expression in hAs. Furthermore, treatment of cells with CM-hDPSCs and hMSCs blocked OGD-induced ROS production and interleukin-1ß upregulation. This study demonstrates for the first time that hDPSCs and CM-hDPSCs confer superior cytoprotection against cell death in an in vitro OGD model compared with hMSCs as shown by cell viability assay. Reactive gliosis, ROS production, and inflammatory mediators might contribute to this protective effect. Therefore, hDPSCs could represent an alternative source of cell therapy for ischemic stroke.

Development of in-cell imaging assay systems for MMP-2 and MMP-9 based on trans-localizing molecular beacon proteins.

A sensitive in-cell imaging MMP-2 and MMP-9 detection systems that enables direct fluorescence detection of a target protease and its inhibition inside living cells has been developed. This in-cell imaging system utilizes the concept of fluorescent molecular beacon reporter (MBR) protein comprising a masking protein, a mitochondrial targeting sequence, a protease specific cleavage sequence and a fluorescent marker sequence, green fluorescent protein (GFP). The MBR protein is designed to change its intracellular location upon cleavage by either MMP-2 or MMP-9 from cytosol to mitochondria. Full and partial MMP-2 and MMP-9 were tested for optimal expression and activity in the cell. The activity of MMP-2 and MMP-9 was approximately 65-71%. Among MMP clones, MMP-2 catalytic domain and MMP-9 clone containing pro, catalytic and hemopexin domain were most active. Both MMP-2 and MMP-9 required divalent ions Ca and Zn for its activity and MMP-9 was more active at higher Ca/Zn ratio. With the in-cell imaging assay the protease activity can be measured in cellular environment and cellular toxicity of candidate molecules can be monitored at the same time. These are great advantage when compared to other currently used in vitro biochemical assays. The in-cell imaging assay developed in this study can be modified for other MMPs and can be used in various life science and drug discovery researches including the high throughput screening and high contents screening applications.