Itraconazole Attenuates Peritoneal Fibrosis Through Its Effect on the Sonic Hedgehog Signaling Pathway in Mice.

BACKGROUND: Peritoneal fibrosis is a devastating complication of peritoneal dialysis. However, its precise mechanism is unclear, and specific treatments have not yet been established. Recent evidence suggests that the sonic hedgehog (SHH) signaling pathway is involved in tissue fibrogenesis. Drugs that inhibit this pathway are emerging in the field of anti-fibrosis therapy. Itraconazole, an anti-fungal agent, was also recently recognized as an inhibitor of the SHH signaling pathway. In this study, we used a mouse model to investigate whether the SHH signaling pathway is involved in the development of peritoneal fibrosis and the effects of itraconazole on peritoneal fibrosis. METHODS: Peritoneal fibrosis was induced by intraperitoneal (IP) injection of 0.1% chlorhexidine gluconate (CG) solution every other day for 4 weeks, with or without itraconazole treatment (20 mg/kg, IP injection on a daily basis). Male C57BL/6 mice were divided into 4 groups: saline group, saline plus itraconazole group, CG group, and CG plus itraconazole group. Isotonic saline was administered intraperitoneally to the control group. The peritoneal tissues were evaluated for histological changes, expression of fibrosis markers, and the main components of the SHH signaling pathway. RESULTS: Peritoneal thickening was evident in the CG group and was significantly decreased by itraconazole administration (80.4 ± 7.7 vs. 28.2 ± 3.8 µm, p < 0.001). The expression of the following SHH signaling pathway components was upregulated in the CG group and suppressed by itraconazole treatment: SHH, patched, smoothened, and glioma-associated oncogene transcription factor 1. The IP injection of CG solution increased the expression of fibrosis markers such as α-smooth muscle actin and transforming growth factor-ß1 in the peritoneal tissues. Itraconazole treatment significantly decreased the expression of these markers. CONCLUSION: Our study provides the first evidence that the SHH signaling pathway may be implicated in peritoneal fibrosis. It also demonstrates that itraconazole treatment has protective effects on peritoneal fibrosis through the regulation of the SHH signaling pathway. These findings suggest that blockage of the SHH signaling pathway is a potential therapeutic strategy for peritoneal fibrosis.

Autophagy activation and neuroprotection by progesterone in the G93A-SOD1 transgenic mouse model of amyotrophic lateral sclerosis.

Progesterone (PG) exerts neuroprotective effects under conditions such as brain ischemia, traumatic brain injury, and spinal cord injury. Previously, we reported that PG activates autophagy, a potential neuroprotective mechanism, in cortical astrocytes. In the present study, we explored the possibility that PG, by activating autophagy in spinal cord cells, protects against motoneuron degeneration in transgenic (Tg) mice expressing the human G93A-SOD1 (superoxide dismutase 1) mutant, a model of amyotrophic lateral sclerosis. PG treatment increased autophagic flux in G93A-SOD1 Tg spinal cord astrocyte cultures and mice. In addition, PG treatment reduced mutant SOD1 protein levels and motoneuronal death. Inhibition of autophagy with 3-methyladenine (3MA) reversed these PG effects, indicating that activation of autophagy contributed to the PG neuroprotection. PG effects in vivo were tested by intraperitoneally injecting male G93A-SOD1 Tg mice with different doses of PG (2, 4, or 8mg/kg) or vehicle from 70days of age until death. Measurements of motor functions using rota-rod tests showed that the onset of symptoms was not different among groups, but the progression of motor dysfunction was significantly delayed in the PG-treated group compared with the vehicle control group. The average lifespan was also prolonged in the PG-injected group. Histological examinations revealed that PG treatment substantially reduced the death of spinal motoneurons at 14weeks of age with a concomitant decrease in mutant SOD1 levels. Our results demonstrated that PG delays neurodegenerative progress in G93A-SOD1 transgenic mice, possibly through activation of autophagy in the spinal cord.

Role of zinc metallothionein-3 (ZnMt3) in epidermal growth factor (EGF)-induced c-Abl protein activation and actin polymerization in cultured astrocytes.

Recent evidence indicates that zinc plays a major role in neurochemistry. Of the many zinc-binding proteins, metallothionein-3 (Mt3) is regarded as one of the major regulators of cellular zinc in the brain. However, biological functions of Mt3 are not yet well characterized. Recently, we found that lysosomal dysfunction in metallothionein-3 (Mt3)-null astrocytes involves down-regulation of c-Abl. In this study, we investigated the role of Mt3 in c-Abl activation and actin polymerization in cultured astrocytes following treatment with epidermal growth factor (EGF). Compared with wild-type (WT) astrocytes, Mt3-null cells exhibited a substantial reduction in the activation of c-Abl upon treatment with EGF. Consistent with previous studies, activation of c-Abl by EGF induced dissociation of c-Abl from F-actin. Mt3 added to astrocytic cell lysates bound F-actin, augmented F-actin polymerization, and promoted the dissociation of c-Abl from F-actin, suggesting a possible role for Mt3 in this process. Conversely, Mt3-deficient astrocytes showed significantly reduced dissociation of c-Abl from F-actin following EGF treatment. Experiments using various peptide fragments of Mt3 showed that a fragment containing the N-terminal TCPCP motif (peptide 1) is sufficient for this effect. Removal of zinc from Mt3 or pep1 with tetrakis(2-pyridylmethyl)ethylenediamine abrogated the effect of Mt3 on the association of c-Abl and F-actin, indicating that zinc binding is necessary for this action. These results suggest that ZnMt3 in cultured astrocytes may be a normal component of c-Abl activation in EGF receptor signaling. Hence, modulation of Mt3 levels or distribution may prove to be a useful strategy for controlling cytoskeletal mobilization following EGF stimulation in brain cells.

Effects of a Neonatal Supportive Positioning Training Video Program for Preterm Infants on the Knowledge and Performance of Nurses in Neonatal Intensive Care Units.

PURPOSE: The purpose of this study is to develop and apply a neonatal supportive positioning (NSP) training video program for premature infants, using a position support mat for nurses in neonatal intensive care units (NICUs), and to verify its effect on nurses' performance. METHODS: Thirty-five NICU nurses were included in the study. For the pre-test, preliminary check-ups were conducted, questionnaires about NSP knowledge on preterm infants were distributed, and NSP performance using neonatal dolls were video recorded for each participant. PowerPoint presentations and videos were used to educate participants on NSP. Furthermore, a 20-minute one-on-one training session was conducted using an NPS kit. Two weeks after the training, we repeated the process of distributing questionnaires about NSP knowledge and recording nurses' performance videos using neonatal dolls. Questionnaires and videos collected before and after the training were compared. RESULTS: After NSP training, the mean knowledge score of the participants improved significantly from 23.71 ± 3.62 to 29.51 ± 2.29 (Z = -5.09, p < .001). The performance score for postural supportive positioning was 38.03 ± 7.46 before training and 80.06 ± 9.85 after receiving training, indicating a high-performance score after NSP training (Z = -5.16, p < .001). CONCLUSION: Our NSP training video program increased nurses' NSP knowledge and performance. Continuous training NICU nurses on NSP, using a standardized training video program, can help improve the care of premature infants.

Effect of Oral Health Education Using a Mobile App (OHEMA) on the Oral Health and Swallowing-Related Quality of Life in Community-Based Integrated Care of the Elderly: A Randomized Clinical Trial.

This study investigated the effect of oral health education using a mobile app (OHEMA) on the oral health and swallowing-related quality of life (SWAL-QoL) of the elderly population in a community-based integrated care project (CICP). Forty elderly individuals in the CICP were randomized into intervention and control groups. OHEMA provided information on customized oral health care management, oral exercises, and intraoral and extraoral massage methods for 50 min/session, once a week, for 6 weeks. Pre- and post-intervention surveys assessed the unstimulated salivary flow rate, subjective oral dryness, tongue pressure, and SWAL-QoL, which were analyzed using ANCOVA and repeated measures ANOVA. In the intervention group, tongue pressure increased significantly from pre- (17.75) to post-intervention (27.24) (p < 0.001), and subjective oral dryness decreased from pre- (30.75) to post-intervention (18.50). The unstimulated salivary flow rate had a higher mean score in the intervention group (7.19) than in the control group (5.04) (p < 0.001). The SWAL-QoL significantly improved from pre- (152.10) to post-intervention (171.50) in the intervention group (p < 0.001) but did not change significantly in the control group (p > 0.05). OHEMA appears to be a useful tool for oral health education for the elderly as it improved the SWAL-QoL, with increased tongue pressure and reduced oral dryness.

Latent tuberculosis infection screening and treatment in congregate settings (TB FREE COREA): protocol for a prospective observational study in Korea.

INTRODUCTION: South Korea regards tuberculosis (TB) incidence in congregate settings as a serious problem. To this end, systematic latent TB infection (LTBI) diagnosis and treatment were provided to approximately 1.2 million individuals in high-risk congregate settings. METHODS AND ANALYSIS: We designed a prospective cohort study of individuals tested for LTBI, based on the data collected on all persons screened for LTBI as part of the 2017 congregate settings programme in South Korea. Four types of databases are kept: LTBI screening database (personal information and LTBI test results), national health information (NHI) database (socio-demographic data and comorbidities), public healthcare information system (PHIS) database, and the Korean national TB surveillance system database (TB outcomes). Information regarding LTBI treatment at private hospitals and public health centres is collected from NHI and PHIS databases, respectively. The screening data are cleaned, duplicates are removed, and, where appropriate, re-coded to analyse specific exposures and outcomes. The primary objective is to compare the number of active TB cases prevented within 2 years between participants undergoing treatment and not undergoing treatment in the LTBI screening programme in congregate settings. Cascade of care for LTBI diagnosis and treatment will be evaluated among those with a positive LTBI test result. A Cox proportional hazards model will be applied to determine the risk factors for developing active TB. ETHICS AND DISSEMINATION: The protocol is approved by the institutional review boards of Incheon St. Mary's Hospital, the Catholic University of Korea. Study results will be disseminated through peer-reviewed journals and conference presentations. TRIAL REGISTRATION NUMBER: KCT0003905.

Oxidative injury triggers autophagy in astrocytes: the role of endogenous zinc.

We have recently demonstrated that the accumulation of labile zinc in lysosomes during oxidative stress causes lysosomal membrane permeabilization (LMP) in cultured hippocampal neurons. Since autophagy involves fusion of autophagic vacuoles (AVs) with lysosomes, zinc accumulation may start in AVs. In the present study, we examined the role of endogenous zinc in H2O2-induced autophagy and cell death in mouse astrocyte cultures. Live-cell confocal imaging of astrocytes transfected with GFP-LC3 revealed that the number of AVs positive for LC3 (microtubule-associated protein 1 light chain 3) increased following exposure to H2O2 or ferrous chloride (FeCl2). Staining of RFP-LC3-transfected astrocytes with FluoZin-3 indicated that the levels of labile zinc increased in AVs as well as in the cytosol and nuclei. The majority of AVs were double-stained with LysoTracker, indicating that they were fused with lysosomes. Chelation of zinc with tetrakis [2-pyridylmethyl]ethylenediamine (TPEN) decreased the number of AVs in H2O2-treated astrocytes, whereas exposure to zinc increased their number, suggesting that dysregulation of zinc homeostasis is mechanistically linked to autophagy. Unexpectedly, inhibition of autophagy blocked the rise in labile zinc levels. Astrocytic death induced by H2O2) was ccompanied by LMP. Autophagy inhibitors (3-methyladenine, bafilomycin-1) or TPEN attenuated LMP and cell death in astrocytes. These results support the possibility that endogenous zinc plays a key role in autophagy under oxidative stress in astrocytes, and suggest that autophagy is a necessary preceding event for LMP and cell death in oxidative injury.

Ethambutol-induced toxicity is mediated by zinc and lysosomal membrane permeabilization in cultured retinal cells.

Ethambutol, an efficacious antituberculosis agent, can cause irreversible visual loss in a small but significant fraction of patients. However, the mechanism of ocular toxicity remains to be established. We previously reported that ethambutol caused severe vacuole formation in cultured retinal cells, and that the addition of zinc along with ethambutol aggravated vacuole formation whereas addition of the cell-permeable zinc chelator, N,N,N',N'-tetrakis (2-pyridylmethyl) ethylenediamine (TPEN), reduced vacuole formation. To investigate the origin of vacuoles and to obtain an understanding of drug toxicity, we used cultured primary retinal cells from newborn Sprague-Dawley rats and imaged ethambutol-treated cells stained with FluoZin-3, zinc-specific fluorescent dye, under a confocal microscope. Almost all ethambutol-induced vacuoles contained high levels of labile zinc. Double staining with LysoTracker or MitoTracker revealed that almost all zinc-containing vacuoles were lysosomes and not mitochondria. Intracellular zinc chelation with TPEN markedly blocked both vacuole formation and zinc accumulation in the vacuole. Immunocytochemistry with antibodies to lysosomal-associated membrane protein-2 (LAMP-2) and cathepsin D, an acid lysosomal hydrolase, disclosed lysosomal activation after exposure to ethambutol. Immunoblotting after 12 h exposure to ethambutol showed that cathepsin D was released into the cytosol. In addition, cathepsin inhibitors attenuated retinal cell toxicity induced by ethambutol. This is consistent with characteristics of lysosomal membrane permeabilization (LMP). TPEN also inhibited both lysosomal activation and LMP. Thus, accumulation of zinc in lysosomes, and eventual LMP, may be a key mechanism of ethambutol-induced retinal cell death.

Antitumor effects of systemically delivered adenovirus harboring trans-splicing ribozyme in intrahepatic colon cancer mouse model.

PURPOSE: Our previous studies suggested that human telomerase reverse transcriptase (hTERT) RNA-targeting trans-splicing ribozyme could be a useful tool for cancer gene therapy. Here, we investigated whether adenoviruses harboring this ribozyme can be systemically delivered to mice, and whether they selectively mark tumors expressing hTERT and sensitize them to ganciclovir treatments. EXPERIMENTAL DESIGN: We constructed adenoviral vectors containing modified hTERT-targeting trans-splicing ribozyme with downstream reporter gene (Ad-Ribo-LacZ) or suicide gene (Ad-Ribo-HSVtk) driven by a cytomegalovirus promoter. The tumor-specific trans-splicing reaction and the tumor-killing effect of adenoviruses harboring ribozyme were investigated both in vitro and in vivo using mice with intrahepatic colon cancer metastasis via systemic administration. The safety of systemic administration of the viruses was also evaluated. RESULTS: We showed that Ad-Ribo-LacZ, when injected i.v., performs a highly specific trans-splicing reaction on hTERT mRNA and that it selectively marks tumors expressing hTERT in mice. More importantly, i.v. injection of Ad-Ribo-HSVtk plus ganciclovir significantly reduced tumor burden, with minimal liver toxicity, in mice with metastatic liver cancer, compared with the untreated group (P = 0.0009). Moreover, animals receiving Ad-Ribo-HSVtk showed improved survival compared with controls (P < 0.0001). CONCLUSIONS: This study shows that systemically delivered adenovirus harboring trans-splicing ribozyme can recognize cancer-specific transcripts and reprogram them to combat the cancer cells. Use of trans-splicing ribozymes seems to be a potentially useful gene therapy for cancer.

The relationship between press release and newspaper coverage of tobacco-related issues in South Korea.

This study investigates an association between press release and news media response on tobacco-related issues in South Korea. We retrieved 231 tobacco-related newspaper articles from all major dailies throughout the year 2005. In total, 37 press releases on tobacco-related issues and policies published by the Korea Ministry of Health and Welfare were obtained from the Ministry website. Content analysis and appropriate statistical tests were performed. Results from our content analysis suggest that producing more press releases on tobacco-related issues may result in a greater volume of newspaper articles, and that a press release on a new topical issue may effect more intense media coverage. Findings also show that when Korean newspaper articles overall held less favorable views of tobacco-related policies and programs in 2005, taxation was the most frequent theme with a non-positive opinion. Findings from our multivariate logistic regression models imply that a newspaper article with a source press release-especially about a new topical issue-is more likely than an article without a source press release to discuss tobacco-related issues more positively. Our findings suggest that a press release may serve as an effective media strategy for reaching out to the public by disseminating tobacco-control efforts and policies.

Job Analysis of the Nursing Unit Managers of Women's Hospital Using DACUM Analysis.

PURPOSE: To analyze the job of nursing unit managers working at women's hospital, using DACUM (developing a curriculum), DACUM is a method for analyzing job-focused competency. METHODS: This study involved a descriptive survey. A DACUM workshop was held to define women's hospital nursing unit managers' role and identify their duties and tasks. For the workshop, a committee was formed consisting of 5 women's hospital nursing unit managers. Finally, after validation, the developed contents were made into a survey asking about nursing unit manager's duties and tasks. RESULTS: Sixteen duties and 83 tasks were identified on the DACUM chart. The importance, difficulty, and frequency of the tasks were ranked in terms of A, B, and C, with A being the highest degree. Eight tasks received A's all in importance, difficulty, and frequency of performance. The 8 tasks were: 'taking over', 'taking care of seriously ill patients on handover', 'ward rounding', 'analyzing and resolving demands identified during handover and patient tour', 'reporting patient status during rounding', 'promoting breast-feeding', 'uterine contraction, and training for breast-feeding'. The duty with the biggest determinant coefficient (DC) was 'patients complaint management' (DC=7.09). Based on tasks, the one with the biggest DC was 'solving patient and patient guardian's complaints' (DC=7.53), followed by 'making infection control guidelines' (DC=7.5). CONCLUSION: When expanding the nursing staff of the hospital, women's hospitals nursing unit managers also need to use administrative functions as intermediaries to focus on the operation management of the entire hospital rather than direct nursing to suit their role.

Tuberculosis control in the Republic of Korea.

The incidence and mortality rates of tuberculosis (TB) in the Republic of Korea are 77 and 5.2 per 100,000 people, respectively (2016), which are the highest among the member countries of the Organization for Economic Cooperation and Development. Recently, the incidence of TB among teens and individuals in their 20s in the Republic of Korea decreased significantly. The decrease is largely attributed to the TB screening and contact investigation efforts targeting schools over the past few years. However, the incidence of TB among elderly individuals remains high, and it is even increasing compared to that in the past 10 years. Older individuals account for 42% of all TB cases and 82% of TB-related deaths. The success rate of TB treatment in the Republic of Korea has gradually increased due to various programs, such as control of non-compliance, insurance coverage for TB diagnosis and treatment, and TB public-private mix models. This study suggests that policy makers should focus their efforts on policies that prioritize a significant reduction in the incidence of TB based on the 2nd National Strategic Plan for Tuberculosis Control (2018-2022).

An Investigation on the Therapeutic Effect of Thymosin ß4 and Its Expression Levels in Streptozotocin-Induced Diabetic Mice.

Thymosin ß4 (Tß4) treatment was known to show the potential therapeutic effects on diabetic complications. This study was performed to determine if Tß4 expression is changed in both serum and tissues under diabetic conditions and can be a serum biomarker. Type 1 diabetic mice were induced in C57/BL6J mice by intraperitoneal injection of streptozotocin (STZ) at a dose of 50 mg/kg body weight. The mice were sacrificed at 16 weeks after STZ injection. Tissues and plasmas were obtained to determine the expression levels of Tß4 using ELISA, real time RT-PCR, and immunohistochemistry. The average serum glucose level was increased to approximately 400 mg/dL beginning 2 weeks after the five injections of STZ and lasting for at least 13 weeks until sacrifice. The plasma and tissue levels of Tß4 in the age-matched control mice were not significantly different from those of the diabetic mice. In conclusion, the Tß4 expression level in the plasmas and tissues of diabetic mice was not affected by diabetic conditions. It indirectly suggests that the therapeutic effect of Tß4 on diabetic complications is due to its regenerative effects on damaged tissue but not to the changed expression level of Tß4 in plasma and tissues of diabetes.

Insulin increases retinal hemorrhage in mild oxygen-induced retinopathy in the rat: inhibition by riluzole.

PURPOSE: Although hyperglycemia is likely the main stimulus for VEGF induction in diabetic retinopathy (DR), a switch from oral hypoglycemic therapy to parenteral insulin injection, despite producing better glucose control, sometimes paradoxically aggravates DR. The induction of VEGF by insulin, as observed in certain conditions, may be a plausible mechanism for this phenomenon. In the present study, to determine the role of insulin in proliferative diabetic retinopathy, the authors examined whether insulin treatment affected the outcome of oxygen-induced retinopathy (OIR) in rats and whether the anti-amyotrophic lateral sclerosis (ALS) drug riluzole with protein kinase C-inhibiting activity can attenuate the effects of insulin. METHODS: To examine in vivo the effects of insulin, mild OIR was produced in 7-day-old rat pups by raising them with a nursing mother in a 55% oxygen environment for 5 days. After that, rat pups were injected daily with subcutaneous saline or insulin (4 U/d) with or without additional riluzole injection (10 mg/kg/d, intraperitoneally) for 5 days in room air. RESULTS: Insulin treatment substantially increased VEGF levels, extraretinal vessel formation, matrix metalloproteinase activity, and the extent of retinal hemorrhage in rat pups with mild OIR compared with saline controls. Riluzole substantially reduced all these changes induced by insulin. CONCLUSIONS: In the present study, OIR was used as a surrogate model for DR because the core pathology and the VEGF-mediated mechanism are shared by both conditions. As in human DR, in rat pups with mild OIR, insulin treatment aggravated retinal hemorrhage, which was blocked by riluzole. Riluzole is a Food and Drug Administration-approved anti-ALS drug with a favorable adverse effect profile. It may be useful as an anti-VEGF treatment in DR, especially in reducing the retinal hemorrhage that often occurs shortly after the switch from oral hypoglycemics to parenteral insulin.

Gene expression responses in vivo by human telomerase reverse transcriptase (hTERT)-targeting trans-splicing ribozyme.

A trans-splicing ribozyme which can specifically reprogram human telomerase reverse transcriptase (hTERT) RNA was previously suggested as a useful agent for tumor-targeted gene therapy. In this study, we evaluated in vivo function of the hTERT-targeting trans-splicing ribozymes by employing the molecular analysis of expression level of genes affected by the ribozyme delivery into peritoneal carcinomatosis mice model. To this effect, we constructed adenoviral vector encoding the specific ribozyme. Noticeably, more than four-fold reduction in the level of hTERT RNA was observed in tumor nodules by the systemic infection of the ribozyme-encoding virus. Such hTERT RNA knockdown in vivo induced changes in the global gene expression profile, including the suppression of specific genes associated with anti-apoptosis including bcl2, and genes for angiogenesis and metastasis. In addition, specific trans-splicing reaction with the targeted hTERT RNA took place in the tumors established as peritoneal carcinomatosis in mice by systemic delivery of the ribozyme. In conclusion, this study demonstrates that an hTERT-specific RNA replacement approach using trans-splicing ribozyme represents a potential modality to treat cancer.

[Analysis of review contents of the submitted papers in Journal of Korean Academy of Nursing--focus: the submitted papers in 2003].

PURPOSE: The objective of the study is to analyze the review contents of reviewers for the submitted papers in the Journal of Korean Academy of Nursing in 2003. METHOD: The review contents of the 165 papers were selected 217 papers that were submitted in 2003. Among those 165 papers, the 21 papers belonged to the 'Do not publish' list and 17 papers, 'Revise manuscript and resubmit', list and the 94 papers, 'Publish if revisions are made' list. There are more than two level differences among the four levels of decision in acceptance of publication in 33 papers. RESULTS: The analysis of the review contents for the papers were suggested according to review categories : introduction, method, results, discussion and conclusion. In addition, if papers had more than two levels of review they were rated poor accord or inconsistent. CONCLUSION: For the quality of the academic journal and development of the nursing science, it is important to disseminate and publish the research paper. Therefore, review of the submitted paper is also important. Implications for the profitable review were suggested in the study.

The zinc ionophore clioquinol reverses autophagy arrest in chloroquine-treated ARPE-19 cells and in APP/mutant presenilin-1-transfected Chinese hamster ovary cells.

Arrested autophagy may contribute to the pathogenesis of Alzheimer's disease. Because we found that chloroquine (CQ) causes arrested autophagy but clioquinol (ClioQ), a zinc ionophore, activates autophagic flux, in the present study, we examined whether ClioQ can overcome arrested autophagy induced by CQ or mutant presenilin-1 (mPS1). CQ induced vacuole formation and cell death in adult retinal pigment epithelial (ARPE-19) cells, but co-treatment with ClioQ attenuated CQ-associated toxicity in a zinc-dependent manner. Increases in lysosome dilation and blockage of autophagic flux by CQ were also markedly attenuated by ClioQ treatment. Interestingly, CQ increased lysosomal pH in amyloid precursor protein (APP)/mPS1-expressing Chinese hamster ovary 7WΔE9 (CHO-7WΔE9) cell line, and ClioQ partially re-acidified lysosomes. Furthermore, accumulation of amyloid-ß (Aß) oligomers in CHO-7WΔE9 cells was markedly attenuated by ClioQ. Moreover, intracellular accumulation of exogenously applied fluorescein isothiocyanate-conjugated Aß(1-42) was also increased by CQ but was returned to control levels by ClioQ. These results suggest that modulation of lysosomal functions by manipulating lysosomal zinc levels may be a useful strategy for clearing intracellular Aß oligomers.

Targeted Regression of Hepatocellular Carcinoma by Cancer-Specific RNA Replacement through MicroRNA Regulation.

Hepatocellular carcinoma (HCC) has a high fatality rate and limited therapeutic options with side effects and low efficacy. Here, we proposed a new anti-HCC approach based on cancer-specific post-transcriptional targeting. To this end, trans-splicing ribozymes from Tetrahymena group I intron were developed, which can specifically induce therapeutic gene activity through HCC-specific replacement of telomerase reverse transcriptase (TERT) RNA. To circumvent side effects due to TERT expression in regenerating liver tissue, liver-specific microRNA-regulated ribozymes were constructed by incorporating complementary binding sites for the hepatocyte-selective microRNA-122a (miR-122a), which is down-regulated in HCC. The ribozyme activity in vivo was assessed in mouse models orthotopically implanted with HCC. Systemic administration of adenovirus encoding the developed ribozymes caused efficient anti-cancer effect and the least hepatotoxicity with regulation of ribozyme expression by miR-122a in both xenografted and syngeneic orthotopic murine model of multifocal HCC. Of note, the ribozyme induced local and systemic antitumor immunity, thereby completely suppressing secondary tumor challenge in the syngeneic mouse. The cancer specific trans-splicing ribozyme system, which mediates tissue-specific microRNA-regulated RNA replacement, provides a clinically relevant, safe, and efficient strategy for HCC treatment.