An Engineered Gene Nanovehicle Developed for Smart Gene Therapy to Selectively Inhibit Smooth Muscle Cells: An In Vitro Study.

In-stent restenosis is a serious concern for patients treated through the stenting procedure, although this can be solved using drug-eluting stents and/or drug-eluting balloon catheters. However, the chemical agents released from the drug-eluting layer for inhibiting smooth muscle cell (SMC) migration are inevitably associated with damage to vascular endothelial cell (ECs). The present in vitro study used a distinct strategy, in which a smart gene (phEGR1-PKCδ, an engineered plasmid consists of an SMC-specific promoter (human early growth response 1, hEGR1 promoter) ligated with a gene encoding apoptosis-inducing protein (protein kinase C-delta, PKCδ) was incorporated into a novel gene vehicle (Au cluster-incorporated polyethylenimine/carboxymethyl hexanoyl chitosan, PEI-Au/CHC) to form the PEI-Au/CHC/phEGR1-PKCδ complex, which was proposed for the selective inhibition of SMC proliferation. It was found that the cell viability of SMCs receiving the PEI-Au/CHC/phEGR1-PKCδ complex under simulated inflammation conditions was significantly lower than that of the ECs receiving the same treatment. In addition, the PEI-Au/CHC/phEGR1-PKCδ complex did not demonstrate an inhibitory effect on EC proliferation and migration under simulated inflammation conditions. Finally, the PEI-Au/CHC/phEGR1-PKCδ complexes coated onto a balloon catheter used in percutaneous transluminal coronary angioplasty (PTCA) could be transferred to both the ECs and the SMC layer of Sprague Dawley (SD) rat aortas ex vivo. These preliminary in vitro results suggest that the newly developed approach proposed in the present study might be a potential treatment for reducing the incidence rate of in-stent restenosis and late thrombosis in the future.

Venlafaxine, paroxetine and milnacipran for major depressive disorder: a pragmatic 24-week study.

Major depressive disorder (MDD), one of the most common psychiatric disorders in the world, is a serious, recurrent and chronic mental disorder, which is associated with significant psychosocial disability and economic burden. Until recently, short-term effectiveness of antidepressants has been measured in terms of patients' response to the medications in significantly reduced depressive symptoms. Remission, a long-term elimination of symptoms and the restoration of normal functioning, has become the primary outcome of therapy. In the current study, the efficacy of three frequently prescribed antidepressants, venlafaxine (75-225 mg/day), paroxetine (20 mg/day) and milnacipran (100 mg/day), used in treating 249 MDD patients with Hamilton Rating Scale of Depression (HRSD17) scores higher than 16 was compared. Each patient was evaluated at week 0, 1, 2, 4, 8, 12, 16, 20 and 24 in a 24-week open-label study. Eighty-two patients took venlafaxine, 97 took paroxetine and 70 patients took milnacipran. No significant differences were found between the three groups in the response condition (HRSD17 scores decreased more than 50%) after 24 weeks of follow-up. For remission, the paroxetine was the least efficacious medication than either the milnacipran (HRSD17 ≤ 7) or the venlafaxine (HRSD17 ≤ 5) by the last observation carried forward (LOCF) analysis. Our results suggest that the absence of depressive symptoms alone may not be an indicator for MDD remission, but the duration of absent depressive symptoms may be a better indicator.

Hemodynamic assessment of the development of portal-systemic collaterals in portal hypertensive rats.

BACKGROUND: Portal hypertension induced the formation of portal-systemic collaterals. Revealing the developmental change of portal-systemic collaterals is important for future therapy. METHODS: We observed the evolution of an accessible shunting vessel, the spleno-renal shunt (SRS), in rats after induction of portal hypertension by partial portal vein ligation (PVL). The hemodynamic and histological assessments of SRS were performed by transit time ultrasound and immunohistochemical studies. RESULTS: The portal pressure of PVL rats surged to 16.5 ± 1.1 mmHg on day 1 after ligation surgery and was maintained at a significantly higher level (13.0 ± 0.4 mmHg) to day 14 when compared to sham rats (p < 0.05). The size or flow of SRS in PVL rats did not change immediately after portal pressure surge. Instead, they increased rapidly on day 4, peaked on day 7, and stabilized thereafter. The size and flow were greater and the resistance of SRS was lower in PVL rats after day 7 (p < 0.05). The anti-Ki67 immunohistochemical study demonstrated positive staining of endothelium in SRS and negative in portal vein or aorta of PVL rats. In addition, the endothelial cells of SRS were stained positive for CD31 and KLF5. CONCLUSIONS: We concluded that the pressure-induced opening of pre-existing vessels was not the primary underlying mechanism in the formation of SRS. Endothelial proliferating and vascular remodeling process participated actively during the development of SRS. These observations can be used for studying the pathogenesis and developing more effective anti-portal hypertensive therapy in the future.

MMP-9 mRNA as a therapeutic marker in acute and chronic stages of arthritis induced by type II collagen antibody.

BACKGROUND/PURPOSE: Antibodies against type II collagen (anti-CII) are arthritogenic and central to the initiation of the disease. An animal model of collagen type II-specific monoclonal antibody-induced arthritis (CAIA) has been used for the evaluation of various therapeutic effects in rheumatoid arthritis (RA). We aimed to measure the expression of matrix metalloproteinase (MMP)-9 (gelatinase B), tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) in the acute and chronic stages of the CAIA model for application as a therapeutic marker. METHODS: A commercially available antibody cocktail containing four monoclonal anti-type II collagen antibodies were injected into 6 to 8-week-old male BALB/c mice (n=20) and 50 microL lipopolysaccharide was injected 3 days later. The clinical manifestations of RA were recorded and scored at 10 days (acute stage) and 21 days (chronic stage). Then the mice were sacrificed for histologic analysis of the inflamed footpad and gene expression of IL-1beta, TNF-alpha and MMP-9 by ELISA and quantitative polymerase chain reaction amplification. RESULTS: Marked inflammation was found in the limb joints of mice at 10 days. Both IL-1beta and MMP-9 expression played a central role in the inflammatory reaction in the acute stage. The expression level of MMP-9 mRNA remained high in the chronic stage of CAIA, but that of IL-1beta mRNA was unexpectedly negligible; the serum level of TNF-alpha in CAIA was undetectable in the acute stage. The expression level of TNF-alpha mRNA was also lower than IL-1beta and MMP-9 in the acute inflammatory stage. CONCLUSION: The CAIA model is a fast and highly replicable model of RA. MMP-9 and IL-1beta were highly expressed in the acute stage of CAIA. It is suggested the MMP-9 mRNA level is a suitable marker for both acute and chronic stage, whereas IL-1beta is a marker only for the acute stage of the CAIA murine model.

D-cycloserine, sarcosine and D-serine diminish the expression of cocaine-induced conditioned place preference.

Reactivation of cocaine-associated memories plays a critical role in reinstating the cocaine-seeking behavior and causing relapse. Cocaine-induced conditioned place preference (CPP) was used as a behavioral paradigm indicative of cocaine-associated memory and repeated cocaine-free preference tests served as a behavioral procedure to retrieve such a memory in this study. Since D-cycloserine was reported to eradicate drug-associated memories, two other N-methyl-D-aspartate (NMDA) receptor agonists were assessed for their efficacy on facilitating the extinction of cocaine-induced CPP. Although D-cycloserine (30 mg/kg) abolished cocaine (10 mg/kg)-induced CPP, sarcosine (300 and 600 mg/kg) and D-serine (600 mg/kg) diminished the expression of such a cocaine memory. Sarcosine (600 mg/kg) and D-serine (600 mg/kg) did not affect the storage of this cocaine memory. It was of interest to note that D-cycloserine facilitated the extinction of cocaine-induced CPP in a fast and early-onset manner, while sarcosine and D-serine decreased cocaine-induced CPP expression in a delay-onset manner. D-cycloserine (30 mg/kg), D-serine (600 mg/kg) and sarcosine (600 mg/kg) did not affect the consolidation of cocaine (5 mg/kg)-induced CPP. Finally, sarcosine (at 600 mg/kg/day for 3 consecutive days) and D-serine (at 600 mg/kg/day for 3 consecutive days) did not produce observable aversive effect associated with their administration in a conditioned place aversion paradigm. Likewise, a similar dosing regimen of sarcosine or D-serine did not cause evident activity-impairing effect. In addition to D-cycloserine treatment, our results indicate that long-term treatment with D-serine and sarcosine may afford a therapeutic advance in suppressing the expression of cocaine-associated memory.

Balanced and unbalanced reciprocal translocation: an overview of a 30-year experience in a single tertiary medical center in Taiwan.

BACKGROUND: Reciprocal translocation is the most common type of translocation; however, there are only a few studies that address the indications for reciprocal translocation in amniocentesis. Here we share our data, based on 30 years' experience in a single tertiary center, to investigate the rates and indications for amniocentesis in cases of reciprocal translocations. METHODS: A retrospective review of 16,749 pregnant women, who underwent midtrimester amniocentesis between January 1981 and December 2010, was conducted. Seventy-four cases of reciprocal translocation were identified. RESULTS: The percentage of reciprocal translocations in all amniocentesis cases was 0.44% (74/16,749); of these 74 cases, 56 were balanced and 18 unbalanced. De novo abnormality occurred in 23 cases, which constituted 31.1% of all reciprocal translocations. The three major indications for amniocentesis with a diagnosis of reciprocal translocation included advanced maternal age (AMA, 52.7%), a parent with an abnormal karyotype (17.6%), and abnormal biochemical markers in the maternal serum (12.2%). For individual types of reciprocal translocations (balanced and unbalanced), except for the presence of abnormal biochemical markers in maternal serum, both AMA and a parent with an abnormal karyotype were primary indications for amniocentesis. However, the highest percentage of reciprocal translocations in all amniocentesis cases was found in cases involving a parent with an abnormal karyotype (5.16%, 13/252). CONCLUSION: Patients with a parent who carries an abnormal karyotype should be encouraged to undergo amniocentesis in prenatal consultation, since the risk of a diagnosis of reciprocal translocation can be particularly high.

Robertsonian translocations: an overview of a 30-year experience in a single tertiary medical center in Taiwan.

BACKGROUND: Advanced maternal age (AMA) is the most frequent indication for amniocentesis in predicting balanced reciprocal translocations, and abnormal ultrasound findings are indications in predicting unbalanced reciprocal translocations; however, to date, no studies have focused on Robertsonian translocations. METHODS: A retrospective review was conducted on 16,749 pregnant women who underwent midtrimester amniocentesis between January 1981 and December 2010. Robertsonian translocations were identified in 39 cases. RESULTS: The percentage of Robertsonian translocations in all amniocentesis cases was 0.23% (39/16,749); 31 were balanced and eight were unbalanced. De novo abnormality occurred in 17 cases, or in 43.6% of all Robertsonian translocations. The two major indications for amniocentesis with a diagnosis of Robertsonian translocations were AMA (41.0%, n = 16) and a parent with abnormal karyotypes (18.0%, n = 7). The highest percentage of Robertsonian translocations was found in parents with abnormal karyotypes (2.8%, 7/252), but neither of the indications were clearly superior for detecting de novo Robertsonian translocations. CONCLUSION: Although AMA is an indication for amniocentesis in approximately two-fifths of cases with Robertsonian translocations, the indication of parent with abnormal karyotypes was more likely to lead to the detection of non-de novo Robertsonian translocations, suggesting that parents with abnormal karyotypes need careful prenatal consultation.

The role of interferon-γ inducible protein-10 in a mouse model of acute liver injury post induced pluripotent stem cells transplantation.

BACKGROUND: Liver injuries are important medical problems that require effective therapy. Stem cell or hepatocyte transplantation has the potential to restore function of the damaged liver and ameliorate injury. However, the regulatory factors crucial for the repair and regeneration after cell transplantation have not been fully characterized. Our study investigated the effects and the expression of the regulatory factors in mouse models of acute liver injury either transplanted with the induced pluripotent stem cells (iPS) or the hepatocytes that differentiated from iPS cells (iHL). METHODS/PRINCIPAL FINDINGS: Mice received CCl(4) injection and were randomized to receive vehicle, iPS, or iHL transfusions vial tail veins and were observed for 24, 48 or 72 hours. The group of mice with iPS transplantation performed better than the group of mice receiving iHL in reducing the serum alanine aminotransferase, aspartate aminotransferase, and liver necrosis areas at 24 hours after CCl(4) injury. Moreover, iPS significantly increased the numbers of proliferating hepatocytes at 48 hours. Cytokine array identified that chemokine IP-10 could be the potential regulatory factor that ameliorates liver injury. Further studies revealed that iPS secreted IP-10 in vitro and transfusion of iPS increased IP-10 protein and mRNA expressions in the injured livers in vivo. The primary hepatocytes and non-parenchyma cells were isolated from normal and injured livers. Hepatocytes from injured livers that received iPS treatment expressed more IP-10 mRNA than their non-hepatocyte counter-parts. In addition, animal studies revealed that administration of recombinant IP-10 (rIP-10) effectively reduced liver injuries while IP-10-neutralizing antibody attenuated the protective effects of iPS and decreased hepatocyte proliferation. Both iPS and rIP-10 significantly reduced the 72-hour mortality rate in mice that received multiple CCl(4)-injuries. CONCLUSIONS/SIGNIFICANCE: These findings suggested that IP-10 may have an important regulatory role in facilitating the repair and regeneration of injured liver after iPS transplantation.

Neonatal isolation decreases cued fear conditioning and frontal cortical histone 3 lysine 9 methylation in adult female rats.

Early life stress is thought to enhance adult susceptibility to stress and stress-related mood disorders. In this study, fear-potentiated startle was used to model the acquisition of a traumatic event-related memory in female rats experiencing early life stress. Daily 1-hr maternal and sibling separation throughout day 2-9 postpartum (D2-9 PP) caused a decrease in the fear-potentiated startle, but not acoustic startle baseline, in adult female rats. The separation procedure did not affect corticosterone secretion but produced an increase in serum estradiol concentration. Moreover, the separation procedure did not affect histone 3 lysine 9 (H3K9) acetylation but decreased H3K9 mono- and tri-methylation in frontal cortices. Treatment with 5-aza-2'-deoxycytidine (AZA) (5mg/kg at alternative days from D2PP to D9PP or 10mg/kg at D5PP and D9PP), a DNA methylation inhibitor, did not affect the separation-decreased fear-potentiated startle. Treatment with valproic acid (VPA), a histone deacetylase inhibitor, at 3 dosing regimens (300mg/kg at D2-9PP; 100mg/kg at D2-4PP, 200mg/kg at D5-7PP, 300mg/kg at D8-9PP; 100mg/kg at D2-5PP, 200mg/kg at D6-9PP) prior to daily separation reversed such a decrease in fear-potentiated startle. The lowest effective VPA dosing regimen used (100mg/kg at D2-5PP, 200mg/kg at D6-9PP) reversed the separation-decreased H3K9 mono- and tri-methylation in frontal cortices. Eight-day VPA (300mg/kg/day) and AZA (5mg/kg/day) administrations starting at D28PP were ineffective in altering the separation-decreased fear-potentiated startle. We, hereby, suggest that decreased frontal cortical H3K9 mono- and tri-methylation may be involved in early life separation-decreased fear memory of adult rats.

An overview of a 30-year experience with amniocentesis in a single tertiary medical center in Taiwan.

OBJECTIVE: Amniocentesis is a popular and effective prenatal diagnostic tool for chromosomal disorders. It is well-established that the risk of chromosomal abnormalities increases with maternal age; however, other related indications are seldom reported. Herein, we report our 30-year experience with amniocentesis from a single medical center, focusing on the indications and rates of abnormality. MATERIAL AND METHODS: A retrospective review of 16,749 pregnant women in the mid-trimester between January 1981 and December 2010 was conducted. The medical records were analyzed. RESULTS: The indications for amniocentesis were advanced maternal age (≥ 34 years old) (n=10,970, 65.5%), increasing-risk maternal triple-marker Down's screening test (≥ 1/270) (n=2090, 12.5%), history of abnormal offspring birth (n=792, 4.7%), abnormal ultrasound findings (n=484, 2.9%), parent with abnormal karyotype (n=252, 1.5%), family history of chromosomal abnormality (n=183, 1.1%), drug and radiation exposure (n=165), abnormal chorionic villus sampling (CVS) results (n=25), intrauterine fetal death (n=50), and other non-specific causes (n=1662, 9.9%). The rate of abnormality for each indication was 16% in the abnormal CVS group, 12% in the intrauterine fetal death group, 11.5% for parental chromosomal abnormality, 8.7% in the abnormal ultrasound finding group, 3.0% in the increasing-risk maternal triple-marker Down's screening test group, 2.5% in the advanced maternal age group, 1.5% for other non-specific causes, 1.4% for history of abnormal offspring birth, and 1.1% for family history of chromosomal abnormality. CONCLUSIONS: Both parents with abnormal karyotype and abnormal ultrasound findings are indications for which consideration of further amniocentesis is highly recommended.

The protective role of natural phytoalexin resveratrol on inflammation, fibrosis and regeneration in cholestatic liver injury.

Liver injuries can trigger a cascade of inflammatory responses and as a result, initiate the process of hepatic regeneration and fibrogenesis. Resveratrol (RSV) has multiple health-promoting benefits. This study evaluated the potential protective effects and mechanism of RSV as related to cholestatic liver injury. RSV was given (4 mg/kg/day, i.p.) for either 3 days or 7 days after bile duct ligation (BDL) injury. RSV significantly reduced serum ALT, AST but not T-bil on Day 3. At this early stage of injury, RSV significantly reduced TNF-α and IL-6 mRNA and decreased the number of Kupffer cells (CD68(+) ) recruited in the injured liver. RSV decreased hepatic fibrosis and reduced collagen Iα1 and TIMP-1 mRNA on Day 7. At the later stages of injury, RSV increased the number of Ki67(+) hepatocytes indicating that RSV promoted hepatocyte proliferation. Additionally, it resulted in decreased expression of 4-hydroxynonenal and increased expression of the hepatocyte growth factor protein and mRNA in the RSV-treated BDL group. Meanwhile, RSV reduced the mortality rate of BDL mice. In conclusion, RSV attenuated inflammation and reduced Kupffer cells activation. RSV decreased fibrosis and promoted hepatocyte regeneration, which increased the survival of BDL mice. RSV was beneficial for the treatment of cholestatic liver injury.

Systemic treatment with protein synthesis inhibitors attenuates the expression of cocaine memory.

It has been proposed that a memory trace enters a labile phase each time it is retrieved. A reactivated memory relies on de novo protein synthesis to be faithfully reconsolidated and restored. Thus, in theory, a long-lasting and pathological memory associated with drug use may be disrupted by inhibiting its reconsolidation through use of protein synthesis inhibitors administered immediately following the memory retrieval. However, effective and efficient strategies to reactivate drug memory remained elusive. This study was undertaken to examine the effects of systemic cycloheximide and anisomycin treatment on the reconsolidation and maintenance of a reactivated cocaine-conditioned place preference (CPP) in mice using several strategies designed to reactivate the previously acquired memory. We found that anisomycin (50 mg/kg/injection) and cycloheximide (15 mg/kg/injection) administered immediately after the reactivation of cocaine-CPP ameliorated subsequent expression and maintenance of this memory. Likewise, when anisomycin and cycloheximide were administered immediately after additional cocaine and saline conditioning trials, the reactivated memory engendered by those extra training trials was also diminished. However, a similar anisomycin dosing regimen failed to affect subsequent expression of cocaine-CPP when additional cocaine conditioning trial was used in the absence of additional saline trial. Finally, cocaine and saline administration to mice in their home cages with or without anisomycin treatment had no effect on later cocaine-CPP expression. Taken together, these findings suggest that systemic treatment with protein synthesis inhibitors immediately after the reactivation of cocaine-CPP effectively diminished the reconsolidation and maintenance of such a cocaine memory. More importantly, reactivation of cocaine-CPP could be achieved by presentation of cocaine-conditioned cues as well as by administering additional cocaine and saline conditioning trials in a balanced fashion.