Protoplasts have been a useful unicellular system for various molecular biological analyses based on transient expression and single cell analysis using fluorescence-activated cell sorting (FACS), widely used as a powerful method in functional genomics. Despite the versatility of these methods, some limits based on low fluorescence intensity of a flow cytometric analysis (FCA) using protoplasts have been reported. In this study, the chloroplast targeting of fluorescent proteins (FPs) led to an eight-fold increase in fluorescence intensity and a 4.5-fold increase of transfection ratio from 14.7% to 65.7% as compared with their targeting into the cytoplasm. Moreover, the plot data of FCA shows that 83.3% of the K-sGFP population is under the threshold level, regarded as a non-transgenic population with background signals, while 65.7% of the K-sGFP population is spread on overall intervals. To investigate the reason underlying this finding, mRNA/protein levels and transfection efficiency were analyzed, and results suggest that mRNA/protein levels and transfection ratio are not much different between K-sGFP and KR-sGFP. From those results, we hypothesized that the difference of fluorescence intensity is not only derived from cellular events such as molecular level or transfection efficiency. Taken together, we suggest that the translocation of FPs into chloroplasts contributes to the improvement of fluorescence intensity in FCA and, apparently, plays an important role in minimizing the loss of the transfected population. Our study could be usefully applicable for highly sensitive FACS and FCA-investigations of green tissue.
chloroplast; flow cytometric analysis (FCA); protoplast; rice; transit peptide; fluorescence-activated cell sorting (FACS); synthetic green fluorescent protein (sGFP)
Metastatic squamous cell carcinoma from a cancer of unknown primary (CUP) affecting the intrathoracic lymph node is very rare. We reported a case of metastatic squamous cell carcinoma in the hilar and interlobar lymph node from a patient with CUP and reviewed the associated literature. Abnormal mass in the right hilar area was incidentally detected. A chest computed tomography scan showed a 2.5-cm diameter mass in the right hilum that had changed little in size for 3 years. The patient underwent a right pneumonectomy and mediastinal lymph node dissection. A metastatic squamous cell carcinoma in the hilar and interlobar lymph nodes without a primary lung or other lesion was diagnosed. The patient received adjuvant chemotherapy for a diagnosis of T0N1M0 lung cancer.
Neoplams; Unknown Primary; Carcinoma; Squamous Cell
Clostridium difficile (C. difficile) is an important cause of nosocomial diarrhea. Diagnostic methods for detection of C. difficile infection (CDI) are shifting to molecular techniques, which are faster and more sensitive than conventional methods. Although recent advances in these methods have been made in terms of their cost-benefit, ease of use, and turnaround time, anaerobic culture remains an important method for detection of CDI.
Materials and Methods
In efforts to evaluate a novel chromogenic medium for the detection of C. difficile (chromID CD agar), 289 fecal specimens were analyzed using two other culture media of blood agar and cycloserine-cefoxitin-fructose-egg yolk agar while enzyme immunosorbent assay and polymerase chain reaction-based assay were used for toxin detection.
ChromID showed the highest detection rate among the three culture media. Both positive rate and sensitivity were higher from chromID than other culture media. ChromID was better at detecting toxin producing C. difficile at 24 h and showed the highest detection rate at both 24 h and 48 h.
Simultaneous use of toxin assay and anaerobic culture has been considered as the most accurate and sensitive diagnostic approach of CDI. Utilization of a more rapid and sensitive chromogenic medium will aid in the dianogsis of CDI.
Clostridium difficile; chromogenic; culture
TGF-β pathway is being extensively evaluated as a potential therapeutic target. The transforming growth factor-β (TGF-β) signaling pathway has the dual role in both tumor suppression and tumor promotion. To design cancer therapeutics successfully, it is important to understand TGF-β related functional contexts. This review discusses the molecular mechanism of the TGF-β pathway and describes the different ways of tumor suppression and promotion by TGF-β. In the last part of the review, the data on targeting TGF-β pathway for cancer treatment is assessed. The TGF-β inhibitors in pre-clinical studies, and Phase I and II clinical trials are updated.
Transforming growth factor-β (TGF-β); EW-7197; ALK5; Breast cancer; Metastasis
The most appropriate treatment for acute gastric variceal bleeding (GVB) is currently endoscopic gastric variceal obturation (GVO) using Histoacryl®. However, the secondary prophylactic efficacy of beta-blocker (BB) after GVO for the first acute episode of GVB has not yet been established. The secondary prophylactic efficacy of BB after GVO for the first acute episode of GVB was evaluated in this study.
Ninety-three patients at Soonchunhyang University Hospital with acute GVB who received GVO using Histoacryl® were enrolled between June 2001 and March 2010. Among these, 42 patients underwent GVO alone (GVO group) and 51 patients underwent GVO with adjuvant BB therapy (GVO+BB group). This study was intended for patients in whom a desired heart rate was reached. The rates of rebleeding-free survival and overall survival were calculated for the two study groups using Kaplan-Meyer analysis and Cox's proportional-hazards model.
The follow-up period after the initial eradication of gastric varices was 18.14±25.22 months (mean±SD). During the follow-up period, rebleeding occurred in 10 (23.8%) and 21 (41.2%) GVO and GVO+BB patients, respectively, and 39 patients died [23 (54.8%) in the GVO group and 16 (31.4%) in the GVO+BB group]. The mean rebleeding-free survival time did not differ significantly between the GVO and GVO+BB groups (65.40 and 37.40 months, respectively; P=0.774), whereas the mean overall survival time did differ (52.54 and 72.65 months, respectively; P=0.036).
Adjuvant BB therapy after GVO using Histoacryl® for the first acute episode of GVB could decrease the mortality rate relative to GVO alone. However, adjuvant BB therapy afforded no benefit for the secondary prevention of rebleeding in GV.
Gastric varices; Beta-blocker; Secondary prevention
To investigate the suitability of citrus-press cakes, by-products of the juice industry as a source for the whitening agents for cosmetic industry.
Ethylacetate extracts of citrus-press cakes (CCE) were examined for their anti-melanogenic potentials in terms of the inhibition of melanin production and mechanisim of melanogenesis by using Western Blot analysis with tyrosinese, tyrosinase-related protein-1 (TRP-1), TRP2, and microphthalmia-associated transcription factor (MITF) proteins. To apply the topical agents, citrus-press cakes was investigated the safety in human skin cell line. Finally flavonoid analysis of CCE was also determined by HPLC analysis.
Results indicated that CCE were shown to down-regulate melanin content in a dose-dependent pattern. The CCE inhibited tyrosinase, TRP-2, and MITF expressions in a dose-dependent manner. To test the applicability of CCE to human skin, we used MTT assay to assess the cytotoxic effects of CCE on human keratinocyte HaCaT cells. The CCE exhibited low cytotoxicity at 50 µg/mL. Characterization of the citrus-press cakes for flavonoid contents using HPLC showed varied quantity of rutin, narirutin, and hesperidin.
Considering the anti-melanogenic activity and human safety, CCE is considered as a potential anti-melanogenic agent and may be effective for topical application for treating hyperpigmentation disorders.
Citrus-press cakes; Melanin; Melanogenesis; Tyrosinase; MITF; TRP-1; TRP-2
Cold urticaria; Cholinergic urticaria
We present a rare case of microgranular variant acute promyelocytic leukemia (APL) associated with ider(17)(q10)t(15;17)(q22;q12) of an old-age patient. The initial chromosome study showed a 46,XX,del(6)(?q21q25),der(15)t(15;17)(q22;q12),ider(17)(q10)t(15;17)/47,sl,+ider(17)(q10)t(15;17)/46,XX. FISH signals from a dual color dual fusion translocation PML-RARA probe were consistent with the results of conventional cytogenetics. Because of the rarity of ider(17)(q10)t(15;17) in microgranular APL, further studies on both gene dosage effect of this chromosomal abnormality and the influence of ider(17)(q10)t(15;17) on clinical features such as prognosis, survival, and treatment response of APL cases are recommended.
ider(17)(q10)t(15;17); Old-age; Microgranular; Acute promyelocytic leukemia
PCR is widely used for rapidly and accurately detecting Mycobacterium Species. The purpose of this study was to assess the diagnostic performance of three real-time PCR kits and evaluate the concordance with two older PCR methods.
Materials and Methods
Using 128 samples, the five PCR methods were assessed, including an in-house PCR protocol, the COBAS Amplicor MTB, the COBAS TaqMan MTB, the AdvanSure TB/NTM real-time PCR, and the Real-Q M. tuberculosis kit. The discrepant results were further examined by DNA sequencing and using the AdvanSure Mycobacteria Genotyping Chip for complete analysis.
For Mycobacterium tuberculosis (MTB) detection, all five kits showed 100% matching results (positive; N = 11 and negative; N = 80). In non-tuberculous mycobacterium (NTM) discrimination, the AdvanSure yielded two true-positive outcomes from M. intracellulare and one false positive outcome, while the Real-Q resulted in one true-positive outcome and one false negative outcome for each case and another false negative result using the provided DNA samples.
Real-time PCR, yielded results that were comparable to those of the older PCR methods for detecting MTB. However, there were disagreements among the applied kits in regard to the sample test results for detecting NTM. Therefore, we recommend that additional confirmatory measures such as DNA sequencing should be implemented in such cases, and further research with using a larger numbers of samples is warranted to improve the detection of NTM.
Mycobacterium tuberculosis; non-tuberculous mycobacterium; real-time PCR
Seaweed has been used in traditional cosmetics and as a herbal medicine in treatments for cough, boils, goiters, stomach ailments, and urinary diseases, and for reducing the incidence of tumors, ulcers, and headaches. Despite the fact that seaweeds are frequently used in the practice of human health, little is known about the role of seaweed in the context of inflammation. This study aimed to investigate the influence of Jeju endemic seaweed on a mouse macrophage cell line (RAW 264.7) under the stimulation of lipopolysaccharide (LPS). Ethyl acetate extracts obtained from 14 different kinds of Jeju seaweeds were screened for inhibitory effects on pro-inflammatory mediators. Our results revealed that extracts from five seaweeds, Laurencia okamurae, Grateloupia elliptica, Sargassum thunbergii, Gloiopeltis furcata, and Hizikia fusiformis, were potent inhibitors of the production of pro-inflammatory mediators such as nitric oxide (NO), prostaglandin E2 (PGE2), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). Based on these results, the anti-inflammatory effects and low cell toxicity of these seaweed extracts suggest potential therapeutic applications in the regulation of the inflammatory response.
Nitric oxide; Interleukin-6 (IL-6); Prostaglandin E2 (PGE2); Tumor necrosis factor-α (TNF-α); Seaweeds; Pro-inflammatory mediators