Small extracellular vesicle-mediated CRISPR-Cas9 RNP delivery for cardiac-specific genome editing.

Myocardial infarction (MI) is a major cause of morbidity and mortality worldwide. Although clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) gene editing holds immense potential for genetic manipulation, its clinical application is hindered by the absence of an efficient heart-targeted drug delivery system. Herein, we developed CRISPR-Cas9 ribonucleoprotein (RNP)-loaded extracellular vesicles (EVs) conjugated with cardiac-targeting peptide (T) for precise cardiac-specific genome editing. RNP complexes containing Cas9 and single guide RNA targeting miR-34a, an MI-associated molecular target, were loaded into EVs (EV@RNP). Gene editing by EV@RNP attenuated hydrogen peroxide-induced apoptosis in cardiomyocytes via miR-34a inhibition, evidenced by increased B-cell lymphoma 2 levels, decreased Bcl-2-associated X protein levels, and the cleavage of caspase-3. Additionally, to improve cardiac targeting in vivo, we used click chemistry to form functional T-EV@RNP by conjugating T peptides to EV@RNP. Consequently, T-EV@RNP-mediated miR-34a genome editing might exert a protective effect against MI, reducing apoptosis, ameliorating MI injury, and facilitating the recovery of cardiac function. In conclusion, the genome editing delivery system established by loading CRISPR/Cas9 RNP with cardiac-targeting EVs is a powerful approach for precise and tissue-specific gene therapy for cardiovascular disease.

Systematic Multiomic Analysis of PKHD1L1 Gene Expression and Its Role as a Predicting Biomarker for Immune Cell Infiltration in Skin Cutaneous Melanoma and Lung Adenocarcinoma.

The identification of genetic factors that regulate the cancer immune microenvironment is important for understanding the mechanism of tumor progression and establishing an effective treatment strategy. Polycystic kidney and hepatic disease 1-like protein 1 (PKHD1L1) is a large transmembrane protein that is highly expressed in immune cells; however, its association with tumor progression remains unclear. Here, we systematically analyzed the clinical relevance of PKHD1L1 in the tumor microenvironment in multiple cancer types using various bioinformatic tools. We found that the PKHD1L1 mRNA expression levels were significantly lower in skin cutaneous melanoma (SKCM) and lung adenocarcinoma (LUAD) than in normal tissues. The decreased expression of PKHD1L1 was significantly associated with unfavorable overall survival (OS) in SKCM and LUAD. Additionally, PKHD1L1 expression was positively correlated with the levels of infiltrating B cells, cluster of differentiation (CD)-8+ T cells, and natural killer (NK) cells, suggesting that the infiltration of immune cells could be associated with a good prognosis due to increased PKHD1L1 expression. Gene ontology (GO) analysis also revealed the relationship between PKHD1L1-co-altered genes and the activation of lymphocytes, including B and T cells. Collectively, this study shows that PKHD1L1 expression is positively correlated with a good prognosis via the induction of immune infiltration, suggesting that PKHD1L1 has potential prognostic value in SKCM and LUAD.

Clinical characteristics and survival of patients concurrently diagnosed with lung cancer and active pulmonary tuberculosis.

Background: The diagnosis of pulmonary tuberculosis (TB) in patients suspected of lung cancer is difficult because of the similarities in signs, symptoms, and radiologic results. The clinical and radiologic characteristics of the co-occurrence of pulmonary TB and lung cancer have not been fully evaluated. Methods: Patients diagnosed with lung cancer and active pulmonary TB from January 2009 to December 2017 in four hospitals of the Catholic University of Korea were retrospectively reviewed. The clinical characteristics, including the TB diagnosis methods, lung cancer pathology, staging, initial radiographic features, and survival were analyzed and compared to 575 lung cancer patients without active pulmonary TB from the same hospitals. Results: Forty-eight (0.48%) of the 9,936 lung cancer patients had active pulmonary TB confirmed for M. tuberculosis at the time of the initial cancer diagnosis. The majority of the patients (95.9%) had non-small cell lung cancer and the most frequent findings were a mass-like lesion (79.2%) and separate nodules (75%). When compared to lung cancer patients without TB, the body mass index (BMI) was lower (21.4 vs. 23.1, P=0.001) and clinical stages were advanced in the lung cancer patients with TB; T3-4 (70.9% vs. 50.6%, P=0.002), N2-3 (85.2% vs. 55.6%, P<0.001); M1 (65.9% vs. 44.5%, P=0.007). Interestingly, lung cancer with TB was associated with lower mortality [hazard ratio (HR) =0.35, 95% CI: 0.21-0.60]. Conclusions: Rarely diagnosed concurrent active TB in lung cancer patients was associated with lower BMI and advanced clinical stages. Active investigation of and treatment for active pulmonary TB in lung cancer patients might possibly improve patient outcomes.

Generation of three TTN knock-out human induced pluripotent stem cell lines using CRISPR/Cas9 system.

TTN mutations are the common genetic cause for various types of cardiomyopathies (e.g., dilated cardiomyopathy, hypertrophic cardiomyopathy, restrictive cardiomyopathy, and arrhythmogenic right ventricular cardiomyopathy) and skeletal myopathies. Here, we generated three TTN knock-out human induced pluripotent stem cell (iPSC) lines using CRISPR/Cas9 system. These cell lines, which exhibit normal karyotype, typical morphology and pluripotency, could provide useful platform for investigating the role of TTN in associated disorders.

Time delays and risk factors in the management of patients with active pulmonary tuberculosis: nationwide cohort study.

Estimating the time delay and identifying associated factors is essential for effective tuberculosis control. We systemically analysed data obtained from the Korea Tuberculosis Cohort in 2019 by classifying delays as presentation and healthcare delays of pulmonary tuberculosis (PTB). Of 6593 patients with active PTB, presentation and healthcare delays were recorded in 4151 and 5571 patients, respectively. The median presentation delay was 16.0 (5.0-40.0) days. Multivariable logistic regression analysis showed that longer presentation delays were associated with neuropsychiatric disease [adjusted odds ratio (OR) 2.098; 95% confidence interval (CI) 1.639-2.687; p < 0.001] and heavy alcohol intake (adjusted OR 1.505; 95% CI 1.187-1.907; p < 0.001). The median healthcare delay was 5.0 (1.0-14.0) days. A longer healthcare delay was associated with malignancy (adjusted OR 1.351; 95% CI 1.069-1.709; p = 0.012), autoimmune disease (adjusted OR 2.445; 95% CI 1.295-4.617; p = 0.006), and low bacterial burden manifested as an acid-fast bacillus smear-negative and tuberculosis polymerase chain reaction-negative status (adjusted OR 1.316; 95% CI 1.104-1.569; p = 0.002). Active case-finding programmes need to focus on patients with heavy alcoholism or neuropsychiatric diseases. To ensure early PTB detection, healthcare providers must carefully monitor patients with malignancy, autoimmune disease, or a high index of suspicion for PTB.

Clinical relevance of false-negative interferon-gamma release assays in patients with tuberculous pleurisy in an intermediate tuberculosis burden country.

Background: There have been few studies to verify factors associated with a false-negative interferon-gamma release assay (IGRA) in patients with tuberculous pleurisy. We investigated the clinical relevance of false-negative results of the blood QuantiFERON-TB Gold In-Tube (QFT-GIT) assay and its risk factors in patients diagnosed with pleural tuberculosis (TB). Methods: Medical records of 650 pleural TB patients in a tertiary hospital between January 2009 and December 2020 were reviewed retrospectively. Patients who underwent the blood QFT-GIT assay and pleural fluid analysis before starting anti-TB medication were included. Results: Of 199 patients with pleural TB who were performed QFT-GIT assay, 36 (18.1%) were false-negative results. These patients tended to be older than those with a positive result (P=0.060). The QFT-GIT-false-negative group of had significantly more comorbidities such as end-stage renal disease (ESRD), haematological cancer or pneumoconiosis than the QFT-GIT-positive group. Hypoproteinaemia and pH >6 in pleural fluid were associated with a false-negative QFT-GIT. Of the 199 patients, 163 (81.9%) were cured or completed anti-TB treatment; 13 patients (6.5%) died. The QFT-GIT-negative patients had significantly worse outcomes including mortality [unfavourable outcome: 33.3% (12/36 patients) in QFT-GIT-negative groups vs. 14.7% (24/163 patients) in QFT-GIT-positive groups, P<0.017; overall mortality: 16.7% (6/36 patients) vs. 4.3% (7/163 patients), respectively, P<0.015]. Conclusions: In pleural TB, a false-negative QFT-GIT result was 18.1% in a country of intermediate TB incidence. This discordant result in GFT-GIT was associated with ESRD, pneumoconiosis, hypoproteinaemia and a poor outcome. Clinicians should keep in mind the possibility of false-negativity in the blood IGRA test, especially in specific situations and its impact on TB outcome in managing patients with pleural TB.

Prognostic Value of Interleukin-32 Expression and Its Correlation with the Infiltration of Natural Killer Cells in Cutaneous Melanoma.

Interleukin-32 (IL-32) is well known as a proinflammatory cytokine that is expressed in various immune cells and cancers. However, the clinical relevance of IL-32 expression in cutaneous melanoma has not been comprehensively studied. Here, we identified the prognostic value of IL32 expression using various systematic multiomic analyses. The IL32 expressions were significantly higher in cutaneous melanoma than in normal tissue, and Kaplan-Meier survival analysis showed a correlation between IL32 expression and good prognosis in cutaneous melanoma patients. In addition, we analyzed the correlation between IL32 expression and the infiltration of natural killer (NK) cells to identify a relevant mechanism between IL32 expression and prognosis in cutaneous melanoma (p = 0.00031). In the relationship between IL32 expression and the infiltration of NK cells, a negative correlation was found in resting NK cells (rho = -0.38, p = 3.95 × 10-17) whereas a strong positive correlation was observed only in active NK cells (rho = 0.374, p = 1.23 × 10-16). Moreover, IL32 expression was markedly positively correlated with the cytolytic molecules, such as granzyme and perforin. These data suggest that IL32 expression may increase patient survival through the infiltration and activation of NK cells, representative anticancer effector cells, in cutaneous melanoma. Collectively, this study provides the prognostic value of IL32 expression and its potential role as an effective predictive biomarker for NK cell infiltration in cutaneous melanoma.

Latent Tuberculosis Infection Screening and Treatment in Congregate Settings (TB FREE COREA): Demographic Profiles of Interferon-Gamma Release Assay Cohort.

In 2017, the Korean government launched an unprecedentedly large-scaled latent tuberculosis infection (LTBI) screening project which covered more than a million individuals in congregate settings. A total of 1,047,689 participants of source population (n = 2,336,157) underwent LTBI testing from 2017 to 2018. The overall LTBI test uptake rate during this project was 44.8%. Workers in daycare centers (83.5%) and kindergartens (78.9%) showed high participation rate. A total of 1,012,206 individuals with valid results of interferon-gamma release assay (IGRA) were selected to constitute the IGRA cohort. Most of the enrolled participants in the IGRA cohort were in their working age. Approximately, three-quarters of total enrolled population were female. Investigating the LTBI prevalence, stages of LTBI care cascade, natural history of LTBI, efficacy of LTBI treatment and cost-effectiveness of LTBI screening are feasible within this IGRA cohort.

Effects of Chronic Low-Dose Internal Radiation on Immune-Stimulatory Responses in Mice.

The Linear-No-Threshold (LNT) model predicts a dose-dependent linear increase in cancer risk. This has been supported by biological and epidemiological studies at high-dose exposures. However, at low-doses (LDR ≤ 0.1 Gy), the effects are more elusive and demonstrate a deviation from linearity. In this study, the effects of LDR on the development and progression of mammary cancer in FVB/N-Tg(MMTVneu)202Mul/J mice were investigated. Animals were chronically exposed to total doses of 10, 100, and 2000 mGy via tritiated drinking water, and were assessed at 3.5, 6, and 8 months of age. Results indicated an increased proportion of NK cells in various organs of LDR exposed mice. LDR significantly influenced NK and T cell function and activation, despite diminishing cell proliferation. Notably, the expression of NKG2D receptor on NK cells was dramatically reduced at 3.5 months but was upregulated at later time-points, while the expression of NKG2D ligand followed the opposite trend, with an increase at 3.5 months and a decrease thereafter. No noticeable impact was observed on mammary cancer development, as measured by tumor load. Our results demonstrated that LDR significantly influenced the proportion, proliferation, activation, and function of immune cells. Importantly, to the best of our knowledge, this is the first report demonstrating that LDR modulates the cross-talk between the NKG2D receptor and its ligands.

Clinical significance of microbial colonization identified by initial bronchoscopy in patients with lung cancer requiring chemotherapy.

BACKGROUND: There are limited data on the association between bronchial colonization and respiratory infections in people with lung cancer requiring cytotoxic chemotherapy. We investigated whether bronchial colonization in initial bronchoscopy specimens can predict the development of pneumonia after chemotherapy in patients with lung cancer. METHODS: Four hundred thirteen patients with lung cancer included in the Catholic Medical Center lung cancer registry were enrolled from March 2015 to August 2018. Demographic data, microbiology results, development of pneumonia after chemotherapy, and clinical information about lung cancer were analyzed retrospectively. RESULTS: A total of 206 lung cancer patients treated with chemotherapy were included in the analysis. Forty patients (19.4%) had positive results for the bronchial washing culture during the initial evaluation of lung cancer. The most common organisms were Klebsiella pneumoniae (n=14) and Streptococcus pneumoniae (n=6) in the surveillance culture, and Pneumocystis jirovecii (n=12) and Staphylococcus aureus (n=8) at the time of pneumonia development. Eighty-nine patients (43.2%) had pneumonia after chemotherapy, but the occurrence of pneumonia did not differ according to the colonization. There were no patients for whom the initial isolated organism was a causative microbe for the development of pneumonia after or during chemotherapy. The pneumonia group had poorer prognosis than the non-pneumonia group (378 vs. 705 days, P=0.0004). CONCLUSIONS: Microbial colonization in bronchoscopy specimens was not associated with pneumonia development or mortality after chemotherapy for lung cancer. This finding suggests that testing surveillance culture may not be helpful for predicting pneumonia or improving survival in lung cancer patients with chemotherapy.

MicroRNA-21 Inhibition Suppresses Alveolar M2 Macrophages in an Ovalbumin-Induced Allergic Asthma Mice Model.

PURPOSE: MicroRNA-21 (miR-21) influences the Th2 immune pathway by suppressing the expressions of interleukin (IL)-12 and interferon (IFN)-γ. The effects of miR-21 suppression on alveolar macrophage polarization and airway inflammation are not known. METHODS: BALB/c and miR-21 knockout (KO) mice were sensitized and challenged with ovalbumin (OVA). The anti-miR-21 antagomir was administered to BALB/c mice by intranasal inhalation from the day of OVA sensitization. Changes in cell counts, cytokine levels in bronchoalveolar lavage fluid (BALF), and airway hyperresponsiveness (AHR) were examined. Total, M1, and M2 macrophages were examined in the lung tissues by immunohistochemistry (IHC). M2 macrophages from the OVA mice lung were inhaled into the anti-miR-21 antagomir-treated asthmatic mice. Moreover, the polarization of M0 to M2 macrophages upon IL-4 stimulation was analyzed after anti-miR-21 antagomir transfection. RESULTS: The miR-21 KO mice showed decreases in AHR, total cell and eosinophil counts in BALF, and in the levels of IL-4, IL-5, IL-10, and IL-13. Expression of IL-12 and IFN-γ were increased in the miR-21 KO mice. Peribronchial inflammation and goblet cell dysplasia were significantly decreased in the lung tissues of miR-21 KO OVA mice compared to the wild type OVA mice. IHC for M1, M2, and total macrophage in the lung tissues showed that miR-21 inhalation suppressed alveolar M2 macrophages in KO mice. M2 macrophage inhalation restored AHR and eosinophilic airway inflammation in the miR-21 antagomir-treated mice. Moreover, anti-miR-21 antagomir transfection decreased the expression of M2 markers and increased the expression of M1 markers in M0 macrophages after IL-4 stimulation. CONCLUSIONS: The results suggest that miR-21 antagonism could suppress alveolar M2 macrophage polarization, decreasing not only the Th2 eosinophilic airway inflammation but also AHR and airway remodeling process.

Urban particulate matter regulates tight junction proteins by inducing oxidative stress via the Akt signal pathway in human nasal epithelial cells.

Recent studies have revealed that increased reactive oxidative stress (ROS) induced by particulate matter (PM) affects tight junction (TJ) functions; however, the molecular mechanisms underlying this effect have not been evaluated fully. Cultured human epithelial cells obtained from inferior turbinate tissues were exposed to an urban PM (UPM) standard reference material (SRM 1648a). Intracellular ROS level and expression of proinflammatory cytokines and TJ proteins were examined. Expression level of phosphorylated (p)-Akt, p38, p65 were compared between exposed and unexposed cells. Cells were pretreated with the ROS scavenger N-acetylcysteine (NAC) or Akt inhibitor MK-2206 before exposure to determine whether the changes in cellular ROS and TJ protein expression could be reversed. Exposure to UPM significantly increased ROS levels and inflammatory cytokine expression levels, and decreased expression of TJ proteins zonula occludins (ZO)-1, occludin, claudin-1, and E-cadherin. UPM exposure increased p-Akt, p-p38, and p65 expression levels, and NAC pretreatment reversed these effects. Akt inhibition decreased UPM-induced ROS formation and p38 and p65 protein phosphorylation, and restored the decreased ZO-1 and E-cadherin expression. Akt inhibition and ROS scavenging may provide targets for maintaining epithelial integrity by restoring decreased TJ protein expression during exposure to UPM.

Evaluation of Human MSCs Treatment Frequency on Airway Inflammation in a Mouse Model of Acute Asthma.

BACKGROUND: Studies in experimental models of allergic asthma have shown that mesenchymal stem cells (MSCs) have therapeutic potential for T-helper 2 (TH2) cell-mediated inflammation. However, the mechanisms underlying these therapeutic effects are not fully understood and their safety has not been confirmed. METHODS: Using a mouse model of experimental allergic asthma, we investigated the efficacy of human adipose-derived mesenchymal stem cells (hADSCs) or human bone marrow-derived mesenchymal stem cells (hBMSCs) according to treatment frequency and timing. RESULTS: Ovalbumin (OVA)-sensitized and -challenged mice exhibited airway hyperresponsiveness (AHR), airway inflammation, and significant increases in TH2 cytokine levels. Both double and single human mesenchymal stem cell (hMSC) treatments significantly decreased AHR and bronchoalveolar lavage fluid counts. In addition, single treatment with hMSCs showed significant attenuation of allergic airway inflammation. However, double treatment with hMSCs during OVA -sensitization and -challenge further increased inflammatory cell infiltration, and TH2 cytokine levels. CONCLUSION: The results of treatment with hADSCs or hBMSCs suppresses AHR and airway inflammation. However, double hMSC treatment significantly induces eosinophilic airway inflammation and lung histological changes. Therefore, double hMSC treatment is ineffective against asthma and single injection frequency appears to be more important for the treatment of asthma. These results suggest that hMSC therapy can be used for treatment of asthma patients but that it should be used carefully.

Neuropilin1 Expression Acts as a Prognostic Marker in Stomach Adenocarcinoma by Predicting the Infiltration of Treg Cells and M2 Macrophages.

Neuropilin1 (NRP1) plays a critical role in tumor progression and immune responses. Although the roles of NRP1 in various tumors have been investigated, the clinical relevance of NRP1 expression in stomach adenocarcinoma (STAD) has not been studied. To investigate the use of NRP1 as a prognostic biomarker of STAD, we analyzed NRP1 mRNA expression and its correlation with patient survival and immune cell infiltration using various databases. NRP1 mRNA expression was significantly higher in STAD than normal tissues, and Kaplan-Meier survival analysis showed that NRP1 expression was significantly associated with poor prognosis in patients with STAD. To elucidate the related mechanism, we analyzed the correlation between NRP1 expression and immune cell infiltration level. In particular, the infiltration of immune-suppressive cells, such as regulatory T (Treg) cells and M2 macrophage, was significantly increased by NRP1 expression. In addition, the expression of interleukin (IL)-35, IL-10, and TGF-ß1 was also positively correlated with NRP1 expression, resulting in the immune suppression. Collectively in this study, our integrated analysis using various clinical databases shows that the significant correlation between NRP1 expression and the infiltration of Treg cells and M2 macrophage explains poor prognosis mechanism in STAD, suggesting the clinical relevance of NRP1 expression as a prognostic biomarker for STAD patients.

A case of early diagnosis of pulmonary capillary hemangiomatosis in a worker with exposure to silica.

BACKGROUND: Pulmonary capillary hemangiomatosis (PCH) is a progressive and refractory vascular disease in the lung. Pulmonary hypertension is frequently combined with PCH when capillary proliferation invades to nearby pulmonary vascular systems. It is difficult to differentiate PCH from other diseases such as pulmonary venoocclusive disease and pulmonary arterial hypertension that cause pulmonary hypertension as they frequently overlap. CASE PRESENTATION: A 29-year-old female who had worked at a bathtub factory presented with progressive exertional dyspnea for the past 2 years. Computed tomography revealed centrilobular, diffusely spreading ground-glass opacities sparing subpleural parenchyma with some cystic lesions and air-trapping in both lungs, suggesting a peculiar pattern of interstitial lung disease with airway involvement. There was not any evidence of right heart failure or pulmonary hypertension on echocardiogram, as well as radiography. Microscopic examination of the lung by thoracoscopic resection showed atypical proliferation of capillary channels within alveolar walls and interlobar septa, without invasion of large vessels. CONCLUSION: We experienced a pathologically diagnosed PCH in a young female complaining progressive dyspnea with prior exposure to occupational silica or organic solvent without elevated right ventricular systolic pressure (RVSP) who showed atypical pattern of radiologic findings.

Korean Guidelines for Diagnosis and Management of Interstitial Lung Diseases: Part 3. Idiopathic Nonspecific Interstitial Pneumonia.

Idiopathic nonspecific interstitial pneumonia (NSIP) is one of the varieties of idiopathic interstitial pneumonias. Diagnosis of idiopathic NSIP can be done via multidisciplinary approach in which the clinical, radiologic, and pathologic findings were discussed together and exclude other causes. Clinical manifestations include subacute or chronic dyspnea and cough that last an average of 6 months, most of which occur in non-smoking, middle-aged women. The common findings in thoracic high-resolution computed tomography in NSIP are bilateral reticular opacities, traction bronchiectasis, reduced volume of the lobes, and ground-glass opacity in the lower lungs. These lesions can involve diffuse bilateral lungs or subpleural area. Unlike usual interstitial pneumonia, honeycombing is sparse or absent. Pathology shows diffuse interstitial inflammation and fibrosis which are temporally homogeneous, namely NSIP pattern. Idiopathic NSIP is usually treated with steroid only or combination with immunosuppressive agents such as azathioprine, cyclophosphamide, cyclosporine, and mycophenolate mofetil. Prognosis of idiopathic NSIP is better than idiopathic pulmonary fibrosis. Many studies have reported a 5-year survival rate of more than 70%.

Metabolic health is more closely associated with decrease in lung function than obesity.

OBJECTIVE: Previous studies have evaluated the link between metabolic syndrome and obesity with impaired lung function, however findings have been controversial. We aimed to compare lung function among subjects with different metabolic health and obesity status. METHODS: Total 10,071 participants were evaluated at the Health Promotion Center in Seoul St. Mary's Hospital between January 2012 and December 2014. Being metabolically healthy was defined as having fewer than three of the following risk factors: high blood pressure, high fasting blood glucose, high triglyceride, low high-density lipoprotein cholesterol and abdominal obesity. Obesity status was defined as body mass index (BMI) higher than 25 kg/m2. Analyses of pulmonary function were performed in four groups divided according to metabolic health and obesity: metabolically healthy non-obese (MHNO), metabolically health obese (MHO), metabolically unhealthy non-obese (MUHNO), and metabolically unhealthy obese (MUHO). RESULTS: Metabolically unhealthy subjects were more prone to decreased lung function compared with their metabolically healthy counterparts, regardless of obesity status. When multinomial logistic regression analysis was performed according to quartiles of forced vital capacity (FVC) or forced expiratory volume in 1 second (FEV1) (% pred), after adjusting for age, sex, and smoking status, odds ratio (OR) for the lowest FVC and FEV1 (% pred) quartiles were significantly higher in MUHO subjects (1.788 [95% CI, 1.531-2.089] and 1.603 [95% CI, 1.367-1.881]) and lower in MHO subjects (0.768 [95% CI, 0.654-0.902] and 0.826 [95% CI, 0.700-0.976]) with MHNO group as the reference, when OR for highest FVC and FEV1 quartiles were considered as 1.0. CONCLUSION: Metabolic health is more closely associated with impaired lung function than obesity.

Cardiac-specific delivery by cardiac tissue-targeting peptide-expressing exosomes.

Naturally occurring RNA carriers such as exosomes might be an untapped source of effective delivery vehicles. However, if exosomes are to be exploited for therapeutic applications, they must target specific tissues or cell types to avoid off-target effects. This study evaluated whether genetic modification of exosomes could enhance exosome delivery to heart cells and heart tissue without toxicity. Exosomes expressing cardiac-targeting peptide (CTP)-Lamp2b on the exosomal membrane (CTP-Exo) were generated by introducing vectors encoding CTP-Lamp2b into HEK 293 cells. The expression of CTP-Lamp2b peptide on exosomes was stabilized by attaching glycosylation sequences. Exosomes expressing only Lamp2b on exosomal membranes (CTL-Exo) were generated as a control. The in vitro and in vivo uptake of CTL-Exo and CTP-Exo was evaluated in cell lines and mice. Both exosomes were delivered to HEK 293 and H9C2 cells. The delivery of the exosome was not different between CTP-Exo and CTL-Exo in HEK 293 cells, whereas the delivery of CTP-Exo was 16% greater than that of CTL-Exo in H9C2 cells (P = 0.047). Cell viability was maintained at almost 100% with different dosages of both CTL-Exo and CTP-Exo. Moreover, compared with CTL-Exo, the in vivo delivery of exosomes to the hearts of mice was increased by 15% with CTP-Exo (P = 0.035). The delivery to livers and spleens was not different between the two exosomes. Genetic modification of exosomes by expressing CTP-Lamp2b on the exosomal membrane enhanced exosome delivery to heart cells and the heart tissue. These results suggested that CTP-Exo might be used as a therapeutic tool for heart disease.

Clinical significance of Glasgow Prognostic Score in patients with tuberculous pleurisy.

BACKGROUND: The Glasgow Prognostic Score (GPS) reflects the host systemic inflammatory response and is a validated, independent prognostic factor for various malignancies. We investigated the clinical significance of the GPS in patients with tuberculosis (TB) pleurisy, focusing on treatment outcomes including paradoxical response (PR). METHODS: This was a retrospective study performed between January 2010 and December 2015 in two referral and university hospitals in South Korea, with intermediate incidences of TB. In all, 462 patients with TB pleurisy were registered in the study. The patients were classified into three groups based on GPS score, as follows: (I) GPS of 2, elevated CRP level (>1.0 mg/dL) and hypoalbuminemia (<3.5 g/dL); (II) GPS of 1, elevated CRP level or hypoalbuminemia; and (III) GPS of 0, neither elevated CRP level nor hypoalbuminemia. RESULTS: A total of 367 patients with TB pleurisy were finally included. PR occurred in 102 (27.8%) patients after a mean of 75 days following initiation of anti-TB treatment. The proportion of PR occurrence was significantly lower in the GPS 2 group (P=0.007). Successful treatment outcomes including cure and completion were also significantly lower in the GPS 2 group (P=0.001), while all-cause mortality and TB-specific mortality were higher in the GPS 2 group (P=0.001 and <0.001, respectively). Old age over than 65 years old was an independent predicting factor for high mortality and lower PR occurrence. However, the TB relapse rate was not different among the three GPS groups. CONCLUSIONS: Higher GPS value and elderly age were identified as prognostic factors for poor outcomes in TB pleurisy and as predicting factors for lower PR occurrence. More prospective studies are needed to clarify the utility of GPS in patients with TB pleurisy.

Uliginosibacterium flavum sp. nov. isolated from an artificial lake.

A Gram-negative, facultative anaerobic, rod-shaped, motile by means of a polar flagellum, greenish-yellow-pigmented bacterial strain (designated strain JJ3220T) was isolated from an artificial lake in South Korea and characterized using a polyphasic approach. The 16S rRNA gene sequence of strain JJ3220T indicated that the isolate belongs to the family Rhodocyclaceae, and that it exhibits 96.4% similarity to Uliginosibacterium paludis KBP-13T. The major cellular fatty acids of the novel strain were C14:0, C16:0, and summed feature 3 (C16:1 ω6c and/or C16:1 ω7c). Strain JJ3220T had flexirubin-type pigments. The DNA G+C content of the strain was 62.8%. The major respiratory quinone and major polar lipid of strain JJ3220T were ubiquinone-8 and phosphatidylethanolamine, respectively. Based on the morphological and physiological properties and biochemical evidence presented, it can be concluded that strain JJ3220T represents a novel species of the genus Uliginosibacterium. The type strain Uliginosibacterium flavum is JJ3220T (=KACC 17644T =JCM 19465T).