Activation of the CD200/CD200R1 axis improves cognitive impairment by enhancing hippocampal neurogenesis via suppression of M1 microglial polarization and neuroinflammation in hypoxic-ischemic neonatal rats.

Following hypoxic-ischemic brain damage (HIBD), there is a decline in cognitive function; however, there are no effective treatment strategies for this condition in neonates. This study aimed to evaluate the role of the cluster of differentiation 200 (CD200)/CD200R1 axis in cognitive function following HIBD using an established model of HIBD in postnatal day 7 rats. Western blotting analysis was conducted to evaluate the protein expression levels of CD200, CD200R1, proteins associated with the PI3K/Akt-NF-κB pathway, and inflammatory factors such as TNF-α, IL-1ß, and IL-6 in the hippocampus. Additionally, double-immunofluorescence labeling was utilized to evaluate M1 microglial polarization and neurogenesis in the hippocampus. To assess the learning and memory function of the experimental rats, the Morris water maze (MWM) test was conducted. HIBDleads to a decrease in the expression of CD200 and CD200R1 proteins in the neonatal rat hippocampus, while simultaneously increasing the expression of TNF-α, IL-6, and IL-1ß proteins, ultimately resulting in cognitive impairment. The administration of CD200Fc, a fusion protein of CD200, was found to enhance the expression of p-PI3K and p-Akt, but reduce the expression of p-NF-κB. Additionally, CD200Fc inhibited M1 polarization of microglia, reduced neuroinflammation, improved hippocampal neurogenesis, and mitigated cognitive impairment caused by HIBD in neonatal rats. In contrast, blocking the interaction between CD200 and CD200R1 with the anti-CD200R1 antibody (CD200R1 Ab) exerted the opposite effect. Furthermore, the PI3K specific activator, 740Y-P, significantly increased the expression of p-PI3K and p-Akt, but reduced p-NF-κB expression. It also inhibited M1 polarization of microglia, reduced neuroinflammation, and improved hippocampal neurogenesis and cognitive function in neonatal rats with HIBD. Our findings illustrate that activation of the CD200/CD200R1 axis inhibits the NF-κB-mediated M1 polarization of microglia to improve HIBD-induced cognitive impairment and hippocampal neurogenesis disorder via the PI3K/Akt signaling pathway.

N6-methyladenosine demethylase FTO regulates synaptic and cognitive impairment by destabilizing PTEN mRNA in hypoxic-ischemic neonatal rats.

Hypoxic-ischemic brain damage (HIBD) can result in significant global rates of neonatal death or permanent neurological disability. N6-methyladenosine (m6A) modification of RNA influences fundamental aspects of RNA metabolism, and m6A dysregulation is implicated in various neurological diseases. However, the biological roles and clinical significance of m6A in HIBD remain unclear. We currently evaluated the effect of HIBD on cerebral m6A methylation in RNAs in neonatal rats. The m6A dot blot assay showed a global augmentation in RNA m6A methylation post-HI. Herein, we also report on demethylase FTO, which is markedly downregulated in the hippocampus and is the main factor involved with aberrant m6A modification following HI. By conducting a comprehensive analysis of RNA-seq data and m6A microarray results, we found that transcripts with m6A modifications were more highly expressed overall than transcripts without m6A modifications. The overexpression of FTO resulted in the promotion of Akt/mTOR pathway hyperactivation, while simultaneously inhibiting autophagic function. This is carried out by the demethylation activity of FTO, which selectively demethylates transcripts of phosphatase and tensin homolog (PTEN), thus promoting its degradation and reduced protein expression after HI. Moreover, the synaptic and neurocognitive disorders induced by HI were effectively reversed through the overexpression of FTO in the hippocampus. Cumulatively, these findings demonstrate the functional importance of FTO-dependent hippocampal m6A methylome in cognitive function and provides novel mechanistic insights into the therapeutic potentials of FTO in neonatal HIBD.

Effect of ultrasound-guided femoral nerve block with dexmedetomidine and ropivacaine on postoperative analgesia in patients undergoing total knee arthroplasty: a randomized controlled trial.

Background: Dexmedetomidine, a potent and highly selective α2-adrenoreceptor agonist, has become a popular adjuvant to local anesthetics. This study was designed to investigate the effect of dexmedetomidine with ropivacaine for femoral nerve block on postoperative analgesia after total knee arthroplasty. Methods: Forty-six patients after total knee arthroplasty received ultrasound-guided femoral nerve block with either 0.3% ropivacaine alone (group R) or 0.3% ropivacaine with 0.5 µg/kg dexmedetomidine (group RD). Total 24-h sufentanil consumption, visual analogue scale (VAS) pain scores, frequency of patient-controlled analgesia (PCA) pressed, Ramsay sedation score, the incidence of bradycardia and hypotension, and incidence of postoperative nausea and vomiting (PONV) were recorded. Results: Compared to group R, the total 24-h sufentanil consumption was significantly reduced (110.76 ± 11.56 vs. 99.09 ± 13.31; P<0.05), the VAS scores were lower at 10 and 12 h postoperatively [3(2-3) vs. 2(1-2) and 3(2-3) vs. 2(1-3), respectively; P<0.05], the frequency of PCA pressed was lower at 8-12 and 12-16-h time intervals [(5(3-6) vs. 2(1-3) and 4(3-4) vs. 2(1-3), respectively; P<0.05]. However, there were no differences in Ramsay's sedation score and the incidence of PONV. Also, no patient experienced bradycardia and hypotension. Conclusions: 0.5 µg/kg dexmedetomidine with 0.3% ropivacaine for femoral nerve block significantly decreased the total 24-h sufentanil consumption, prolonged and enhanced the analgesic efficacy of ropivacaine, without clinically relevant cardiovascular depression or over-sedation in patients undergoing total knee arthroplasty.

Activation of the CD200/CD200R1 axis attenuates neuroinflammation and improves postoperative cognitive dysfunction via the PI3K/Akt/NF-κB signaling pathway in aged mice.

BACKGROUND: Postoperative cognitive dysfunction (POCD) is a neurological complication occurring after anesthesia and surgery. Neuroinflammation plays a critical role in the pathogenesis of POCD, and the activation of the cluster of differentiation 200 (CD200)/CD200R1 axis improves neurological recovery in various neurological disorders by modulating inflammation. The aim of this study was to investigate the impact and underlying mechanism of CD200/CD200R1 axis on POCD in aged mice. METHODS: The model of POCD was established in aged mice. To assess the learning and memory abilities of model mice, the Morris water maze test was implemented. CD200Fc (CD200 fusion protein), CD200R1 Ab (anti-CD200R1 antibody), and 740Y-P (a specific PI3K activator) were used to evaluate the effects of the CD200/CD200R1/PI3K/Akt/NF-κB signaling pathway on hippocampal microglial polarization, neuroinflammation, synaptic activity, and cognition in mice. RESULTS: It was observed that anesthesia/surgery induced cognitive decline in aged mice, increased the levels of tumor necrosis factor alpha (TNF-α), interleukin (IL)-6, IL-1 ß and decreased the levels of postsynaptic density protein 95 (PSD-95), synaptophysin (SYN) in the hippocampus. Moreover, CD200Fc and 740Y-P attenuated neuroinflammation and synaptic deficits and reversed cognitive impairment via the phosphatidylinositol 3-kinase (PI3K)/ protein kinase B (Akt)/nuclear factor-kappa B (NF-κB) signaling pathway, whereas CD200R1 Ab administration exerted the opposite effects. Our results further show that the CD200/CD200R1 axis modulates M1/M2 polarization in hippocampal microglia via the PI3K/Akt/NF-κB signaling pathway. CONCLUSIONS: Our findings indicate that the activation of the CD200/CD200R1 axis reduces neuroinflammation, synaptic deficits, and cognitive impairment in the hippocampus of aged mice by regulating microglial M1/M2 polarization via the PI3K/Akt/NF-κB signaling pathway.

Dexmedetomidine Regulates Autophagy via the AMPK/mTOR Pathway to Improve SH-SY5Y-APP Cell Damage Induced by High Glucose.

Neurodegenerative diseases and postoperative cognitive dysfunction involve the accumulation of ß-amyloid peptide (Aß). High glucose can inhibit autophagy, which facilitates intracellular Aß clearance. The α2-adrenoreceptor agonist dexmedetomidine (DEX) can provide neuroprotection against several neurological diseases; however, the mechanism remains unclear. This study investigated whether DEX regulated autophagy via the AMPK/mTOR pathway to improve high glucose-induced neurotoxicity in SH-SY5Y/APP695 cells. SH-SY5Y/APP695 cells were cultured with high glucose with/without DEX. To examine the role of autophagy, the autophagy activator rapamycin (RAPA) and autophagy inhibitor 3-methyladenine (3-MA) were used. The selective AMPK inhibitor compound C was used to investigate the involvement of the AMPK pathway. Cell viability and apoptosis were examined by CCK-8 and annexin V-FITC/PI flow cytometric assays, respectively. Autophagy was analyzed by monodansylcadaverine staining of autophagic vacuoles. Autophagy- and apoptosis-related protein expression and the phosphorylation levels of AMPK/mTOR pathway molecules were quantified by western blotting. DEX pretreatment significantly suppressed high glucose-induced neurotoxicity in SH-SY5Y/APP695 cells, as evidenced by the enhanced viability, restoration of cellular morphology, and reduction in apoptotic cells. Furthermore, RAPA had a protective effect similar to that of DEX, but 3-MA eliminated the protective effect of DEX by promoting mTOR activation. Moreover, the AMPK/mTOR pathway was involved in DEX-mediated autophagy. Compound C significantly suppressed autophagy and reversed the protective effect of DEX against high glucose in SH-SY5Y/APP695 cells. Our findings demonstrated that DEX protected SH-SY5Y/APP695 cells against high glucose-induced neurotoxicity by upregulating autophagy through the AMPK/mTOR pathway, suggesting a role of DEX in treating POCD in diabetic patients.

Dexmedetomidine alleviates olfactory cognitive dysfunction by promoting neurogenesis in the subventricular zone of hypoxic-ischemic neonatal rats.

Background: Hypoxic-ischemic brain damage (HIBD) is the main cause of neurological dysfunction in neonates. Olfactory cognitive function is important for feeding, the ability to detect hazardous situations and social relationships. However, only a few studies have investigated olfactory cognitive dysfunction in neonates with HIBD; furthermore, the specific mechanisms involved are yet to be elucidated. It has been reported that neurogenesis in the subventricular zone (SVZ) is linked to olfactory cognitive function. Recently, dexmedetomidine (DEX) has been shown to provide neuroprotection in neonates following HIBD. In the present study, we investigated whether DEX could improve olfactory cognitive dysfunction in neonatal rats following HIBD and attempted to determine the underlying mechanisms. Methods: We induced HIBD in rats using the Rice-Vannucci model, and DEX (25 µg/kg, i.p.) was administered immediately after the induction of HIBD. Next, we used triphenyl tetrazolium chloride (TTC) staining and the Zea-longa score to assess the success of modelling. The levels of BDNF, TNF-α, IL-1ß and IL-6 were determined by western blotting. Immunofluorescence staining was used to detect microglial activation and microglial M1/M2 polarization as well as to evaluate the extent of neurogenesis in the SVZ. To evaluate the olfactory cognitive function, the rats in each group were raised until post-natal days 28-35; then, we performed the buried food test and the olfactory memory test. Results: Analysis showed that HIBD induced significant brain infarction, neurological deficits, and olfactory cognitive dysfunction. Furthermore, we found that DEX treatment significantly improved olfactory cognitive dysfunction in rat pups with HIBD. DEX treatment also increased the number of newly formed neuroblasts (BrdU/DCX) and neurons (BrdU/NeuN) in the SVZ by increasing the expression of BDNF in rat pups with HIBD. Furthermore, analysis showed that the neurogenic effects of DEX were possibly related to the inhibition of inflammation and the promotion of M1 to M2 conversion in the microglia. Conclusion: Based on the present findings, DEX treatment could improve olfactory cognitive dysfunction in neonatal rats with HIBD by promoting neurogenesis in the SVZ and enhancing the expression of BDNF in the microglia. It was possible associated that DEX inhibited neuroinflammation and promoted M1 to M2 conversion in the microglia.

Visible-Light-Promoted Desulfurative Alkylation of Alkyl Thianthrenium Salts with Activated Olefins.

Reactions involving an alkyl radical generated from a primary alcohol by photochemistry are rare and challenging. Herein, we present a photocatalyst- and metal-free approach that enables the generation of an alkyl radical from the corresponding alcohol and the subsequent C(sp3)-C(sp3) bond formation with activated olefin, via an alkyl thianthrenium salt/Hantzsch ester electron donor-acceptor complex. This protocol for the conversion of a C-OH bond to a C-C bond is highly functionality tolerant and can successfully be used in late-stage functionalization of pharmaceuticals.

Effect of Ultrasound-Guided Fascia Iliac Compartment Block with Nalbuphine and Ropivacaine on Preoperative Pain in Older Patients with Hip Fractures: A Multicenter, Triple-Blinded, Randomized, Controlled Trial.

INTRODUCTION: Pain management for older patients with hip fractures is challenging. This study aimed to investigate the effect of ultrasound-guided fascia iliac compartment block (UGFICB) using different doses of nalbuphine in combination with ropivacaine on preoperative analgesia in older patients with hip fractures. METHODS: In this multicenter randomized controlled trial, 280 elderly patients with hip fracture were randomly allocated into four UGFICB groups (n = 70 in each group): a ropivacaine group (30 mL 0.1% ropivacaine + 0.9% normal saline) and three ropivacaine plus nalbuphine groups (5, 10, and 20 mg nalbuphine, respectively). The primary outcomes were the duration of analgesia at rest and on passive movement. Secondary outcomes included sensory block area, side effects, and vital signs. The doses of rescue analgesia with parecoxib sodium were also analyzed. RESULTS: The addition of nalbuphine dose-dependently increased the duration of analgesia at rest and on passive movement (P < 0.05) and expanded the area of sensory block (P < 0.05). Compared with the ropivacaine group, the pain scores at rest and on movement at 6 and 8 h after the block were lower in three ropivacaine plus nalbuphine groups (P < 0.05), without between-group differences at 2, 4, and 12 h. The four groups had comparable side effects (nausea and vomiting) and vital signs (P > 0.05). CONCLUSIONS: UGFICB with 5, 10, and 20 mg nalbuphine added to ropivacaine prolonged the analgesia duration, increased sensory block area, reduced pain, and decreased the doses of rescue parecoxib sodium for older patients after hip fracture, without obvious side effects. Among these three doses, nalbuphine 20 mg in combination with ropivacaine provided the longest duration of analgesia and the largest sensory block area. TRIAL REGISTRATION: Chinese Clinical Trial Registry (ChiCTR2000029934).

Ultrasound-guided transversus abdominis plane block using ropivacaine and dexmedetomidine in patients undergoing caesarian sections to relieve post-operative analgesia: A randomized controlled clinical trial.

Dexmedetomidine, which is a highly selective α2 adrenoreceptor agonist, enhances the analgesic efficacy and prolongs the analgesic duration when administered in combination with local anesthetics. The current study aimed to evaluate the effects of dexmedetomidine combined with ropivacaine in ultrasound-guided transversus abdominis plane (TAP) block on post-operative analgesia following cesarean section (CS). A total of 70 patients scheduled for CS were divided randomly into 2 groups: The ropivacaine (R) group, in which patients were administered bilateral 20 ml 0.3% ropivacaine and 2 ml 0.9% normal saline, and the dexmedetomidine (RD) group, in which patients were administered bilateral 20 ml 0.3% ropivacaine and 2 ml dexmedetomidine (0.5 µg/kg). The primary outcome was pain-free duration, and secondary outcomes included heart rate (HR) and mean blood pressure (MBP) measurements, visual analogue scale (VAS) pain scores, number of patients who required rescue analgesic, time to first request for analgesia and patient satisfaction. There was no significant difference in HR and MBP between the two groups at 1 h post-surgery (P>0.05). However, VAS pain scores decreased at 6 and 8 h post-surgery [2 (1-2) vs. 0 (0-0.25) and 2 (2-3) vs. 0 (0-1), respectively; P<0.05], pain-free duration was prolonged (5.91±1.08 vs. 9.62±1.46 h; P<0.05), the number of patients who required rescue analgesic was reduced (19 vs. 9; P<0.05), the time to first request for analgesia was prolonged (7.10±1.21 vs. 11.60±2.11 h; P<0.05) and patient satisfaction was improved [3.5 (3-4) vs. 4 (4-5); P<0.05] in the RD group compared with the R group. Furthermore, no bradycardia or hypotension was observed. In conclusion, the results of the present study demonstrated that adding 0.5 µg/kg dexmedetomidine to 0.3% ropivacaine used in TAP block in patients undergoing CS prolonged pain-free duration, decreased VAS pain scores, reduced the number of patients who required rescue analgesic, prolonged the time to first request for analgesia and improved the patient satisfaction without serious side effects.

CD200R Is Involved in the Anti-inflammatory Effect of Dexmedetomidine in Lipopolysaccharide-Stimulated Microglia.

Dexmedetomidine has been confirmed to have anti-inflammatory effect in numerous diseases. However, the detailed mechanism is still unknown. We explore the hypothesis that CD200R is related to the anti-inflammatory effect of dexmedetomidine. Primary microglia were treated with different concentrations of dexmedetomidine followed with lipopolysaccharide (LPS) for 24 h. Interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and interleukin-10 (IL-10) were measured by enzyme-linked immunosorbent assay. Arginase 1 (Arg1), IL-6, and TNF-α were measured by TR-PCR. CD200R was observed by Western blot analysis. We found dexmedetomidine reduced LPS-induced IL-6 and TNF-α production and increase Arg1 in primary microglia. CD200R expression was downregulated by LPS and dexmedetomidine can attenuate the effect of LPS. Our results showed that CD200R may be related to the anti-inflammatory effect of dexmedetomidine.

[Dexmedetomidine alleviates cognitive dysfunction induced by tibial fracture in rats].

OBJECTIVE: To study the effect of dexmedetomidine (Dex) on cognitive dysfunction induced by tibial fracture in rats. METHODS: Sixteen male SD rats were randomized into control group and tibial fracture group, and the behavior indicators were measured on days 1, 3, 5 and 7 after tibial fracture and the expressions of CX3L1 protein and mRNA in the hippocampus were detected. Another 24 male SD rats were randomly divided into control group, tibial fracture group, and tibia fracture with CX3CL1 antibody group, and the behavior indicators and hippocampal CX3L1 protein expression were evaluated after corresponding treatments. In another experiment, we randomized 24 male SD rats into control group, tibial fracture group and tibial fracture with Dex treatment, and tested their hippocampal CX3L1 protein and mRNA expressions as well as the behavior indicators after the treatments. RESULTS: Compared with the control rats, the rats with tibial fracture spent significantly less time in the novel arm on days 1, 3, 5 and 7 after the fracture (P < 0.05) with obviously lowered expressions of CX3L1 protein and mRNA in the hippocampus (P < 0.05). In the rats with tibial fracture, treatment with CX3CL1 antibody further decreased the time spent in the novel arm (P < 0.05) and the expression level of CX3L1 protein in the hippocampus (P < 0.05); In contrast, treatment with Dex significantly increased the time spent time in the novel arm (P < 0.05) and enhanced the hippocampal expressions of CX3L1 protein and mRNA in rats with tibial fractures (P < 0.05). CONCLUSIONS: Dex can alleviate cognitive dysfunction induced by tibial fracture in rats by increasing the expression of CX3CL1 in the hippocampus.

Dexmedetomidine inhibits inflammatory reaction in the hippocampus of septic rats by suppressing NF-κB pathway.

Dexmedetomidine (DEX) is known to provide neuroprotective effect in the central nervous system. However, the detailed mechanism remains far more elusive. This study was designed to investigate the relevant mechanisms of DEX's neuroprotective effect. Sprague-Dawley (SD) rats were injected with dexmedetomidine and/or Lipopolysaccharide (LPS) intraperitoneally, and inflammatory cytokines in serum and in the hippocampus were measured by enzyme linked immunosorbent assay (ELISA). NF-κB in the brain tissue extracts was analyzed with western-blot. Then, we investigated whether NF-κB inhibitor prevents the elevation of inflammatory cytokines in rats injected with LPS. Our results indicated that compared with the control group, the rats exposed to LPS showed significant cognitive dysfunction. When compared to controls, the levels of TNF-α and IL-6 in the serum and hippocampus homogenate were increased in rats treated with LPS. DEX pretreatment inhibited the rats' TNF-α, IL-6 and NF-κB levels induced by LPS. In response to LPS, PDTC pretreatment restrains the production of proinflammatory cytokines (TNF-α and IL-6). Rats treated with PDTC and DEX alongside LPS exhibited less TNF-α and IL-6 than the LPS treated group. In combination, PDTC and DEX showed addictive effects. Our data suggest that DEX exerts a neuroprotective effect through NF-κB in part after LPS-induced cognitive dysfunction.

Anisotropic Failure Strength of Shale with Increasing Confinement: Behaviors, Factors and Mechanism.

Some studies reported that the anisotropic failure strength of shale will be weakened by increasing confinement. In this paper, it is found that there are various types of anisotropic strength behaviors. Four types of anisotropic strength ratio ( S A 1 ) behaviors and three types of anisotropic strength difference ( S A 2 ) behaviors have been classified based on laboratory experiments on nine groups of different shale samples. The cohesion c w and friction angle ϕ w of the weak planes are proven to be two dominant factors according to a series of bonded-particle discrete element modelling analyses. It is observed that shale is more prone to a slight increase of S A 1 and significant increase of S A 2 with increasing confinement for higher cohesion c w and lower to medium friction angle ϕ w . This study also investigated the mechanism of the anisotropic strength behaviors with increasing confinement. Owing to different contributions of c w and ϕ w under different confinements, different combinations of c w and ϕ w may have various types of influences on the minimum failure strength with the increasing confinement; therefore, different types of anisotropic behaviors occur for different shale specimens as the confinement increases. These findings are very important to understand the stability of wellbore and underground tunneling in the shale rock mass, and should be helpful for further studies on hydraulic fracture propagations in the shale reservoir.

Genetic variation and geographic differentiation among populations of the nonmigratory agricultural pest Oedaleus infernalis (Orthoptera: Acridoidea) in China.

The nonmigratory grasshopper Oedaleus infernalis Saussure (Orthoptera : Acridoidea) is an agricultural pest to crops and forage grasses over a wide natural geographical distribution in China. The genetic diversity and genetic variation among 10 geographically separated populations of O. infernalis was assessed using polymerase chain reaction-based molecular markers, including the intersimple sequence repeat and mitochondrial cytochrome oxidase sequences. A high level of genetic diversity was detected among these populations from the intersimple sequence repeat (H: 0.2628, I: 0.4129, Hs: 0.2130) and cytochrome oxidase analyses (Hd: 0.653). There was no obvious geographical structure based on an unweighted pair group method analysis and median-joining network. The values of FST, θ(II), and Gst estimated in this study are low, and the gene flow is high (Nm > 4). Analysis of the molecular variance suggested that most of the genetic variation occurs within populations, whereas only a small variation takes place between populations. No significant correlation was found between the genetic distance and geographical distance. Overall, our results suggest that the geographical distance plays an unimpeded role in the gene flow among O. infernalis populations.

A simple confined impingement jets mixer for flash nanoprecipitation.

Johnson and Prud'homme (2003. AICHE J 49:2264-2282) introduced the confined impingement jets (CIJ) mixer to prepare nanoparticles loaded with hydrophobic compounds (e.g., drugs, inks, fragrances, or pheromones) via flash nanoprecipitation (FNP). We have modified the original CIJ design to allow hand operation, eliminating the need for a syringe pump, and we added a second antisolvent dilution stage. Impingement mixing requires equal flow momentum from two opposing jets, one containing the drug in organic solvent and the other containing an antisolvent, typically water. The subsequent dilution step in the new design allows rapid quenching with high antisolvent concentration that enhances nanoparticle stability. This new CIJ with dilution (CIJ-D) mixer is a simple, cheap, and efficient device to produce nanoparticles. We have made 55 nm diameter ß-carotene nanoparticles using the CIJ-D mixer. They are stable and reproducible in terms of particle size and distribution. We have also compared the performance of our CIJ-D mixer with the vortex mixer, which can operate at unequal flow rates (Liu et al., 2008. Chem Eng Sci 63:2829-2842), to make ß-carotene-containing particles over a series of turbulent conditions. On the basis of dynamic light scattering measurements, the new CIJ-D mixer produces stable particles of a size similar to the vortex mixer. Our CIJ-D design requires less volume and provides an easily operated and inexpensive tool to produce nanoparticles via FNP and to evaluate new nanoparticle formulation.

A Strategy for Control of "Random" Copolymerization of Lactide and Glycolide: Application to Synthesis of PEG-b-PLGA Block Polymers Having Narrow Dispersity.

Poly(lactic-co-glycolic acid) (PLGA) is a biodegradable copolymer that is also acceptable for use in a variety of biomedical applications. Typically, a random PLGA polymer is synthesized in a bulk batch polymerization using a tin-based catalyst at high temperatures. This methodology results in relatively broad polydispersity indexes (PDIs) due to transesterification, and the polymer product is often discolored. We report here the use of 1,8-diazabicyclo[5.4.0]-undec-7-ene (DBU), a known, effective, and convenient organocatalyst for the ring-opening polymerization of cyclic esters, to synthesize random copolymers of lactide and glycolide. The polymerization kinetics of the homo- and copolymerizations of lactide and glycolide were explored via NMR spectroscopy. A novel strategy that employs a controlled addition of the more reactive glycolide monomer to a solution containing the lactide monomer, the poly(ethylene glycol) (PEG) macroinitiator, and DBU catalyst was developed. Using this tactic (semi-batch polymerization), we synthesized a series of block copolymers that exhibited excellent correlation of the expected and observed molecular weights and possessed narrow PDIs. We also measured the thermal properties of these block copolymers and observed trends based on the composition of the block copolymer. We also explored the need for experimental rigor in several aspects of the preparations and have identified a set of convenient reaction conditions that provide polymer products that retain the aforementioned desirable characteristics. These polymerizations proceed rapidly at room temperature and without the need for tin-based catalysts to provide PEG-b-PLGAs suitable for use in biomedical investigations.

[Construction of expression vector of hTERT/hIL-18 fusion gene in eukaryotic cells and its function].

OBJECTIVE: To construct expression vector of hTERT-hIL-18 fusion gene in eukaryotic cells and to study its biological function. METHODS: hIL-18 gene was amplified by RT-PCR, then T-A cloned and inserted into PCDNA3.1(+)/hTERT vector. The sequence of fusion gene was examined by enzyme incision and DNA sequencing. The vector with fusion gene was transformed into 3T3 cells by the method of lipofecting, and proved by Western blot. The secretion gamma-interferon was measured with ELISA and cell apoptosis was detected with flow cytometry. RESULT: Expression vector PCDNA3.1(+) of hTERT/hIL-18 fusion gene was constructed successfully. The correct sequence was proved by enzyme incision and sequencing and there was a correct open reading frame. Fusion protein of hTERT/hIL-18 was effectively expressed in eukaryotic cells and was proved by Western blot and immunofluorescence stain. The fusion protein stimulated KG-1 cells to secrete gamma-interferon and had anti-apoptosis effect. CONCLUSION: Fusion protein hTERT-hIL-18 is highly effectively expressed in eukaryotic cells and is biologically active.

Intraperitoneal stability of alginate-polyornithine microcapsules in rats: an FTIR and SEM analysis.

Alginate-polycation microcapsule systems have been used over decades as delivery vehicles for cell and protein therapy. These systems have been unpredictable across a range of indications with questions resulting around the inherent stability of the alginate polysaccharide and failure mode of the delivery system. The current study focuses on such a system using 5 different alginates, 2 of which are commercially purified, which are crosslinked by polyornithine. Capsules formed by frequency-generated droplet formation were studied in the peritoneal cavity of Long-Evans rats over the course of 3 months by morphometry, Fourier-transform infrared spectroscopy (FTIR), and scanning electron microscopy of the surface. Individual capsule components were also investigated on FTIR and a relative stability index was generated by titration for comparison to explanted samples over time. Using these techniques, a distinct degradation pattern was noted and is compared between the 5 alginate sources.