Current application and future perspective of CRISPR/cas9 gene editing system mediated immune checkpoint for liver cancer treatment.

Liver cancer, which is well-known to us as one of human most prevalent malignancies across the globe, poses a significant risk to live condition and life safety of individuals in every region of the planet. It has been shown that immune checkpoint treatment may enhance survival benefits and make a significant contribution to patient prognosis, which makes it a promising and popular therapeutic option for treating liver cancer at the current time. However, there are only a very few numbers of patients who can benefit from the treatment and there also exist adverse events such as toxic effects and so on, which is still required further research and discussion. Fortunately, the clustered regularly interspaced short palindromic repeat/CRISPR-associated nuclease 9 (CRISPR/Cas9) provides a potential strategy for immunotherapy and immune checkpoint therapy of liver cancer. In this review, we focus on elucidating the fundamentals of the recently developed CRISPR/Cas9 technology as well as the present-day landscape of immune checkpoint treatment which pertains to liver cancer. What's more, we aim to explore the molecular mechanism of immune checkpoint treatment in liver cancer based on CRISPR/Cas9 technology. At last, its encouraging and powerful potential in the future application of the clinic is discussed, along with the issues that already exist and the difficulties that must be overcome. To sum up, our ultimate goal is to create a fresh knowledge that we can utilize this new CRISPR/Cas9 technology for the current popular immune checkpoint therapy to overcome the treatment issues of liver cancer.

Dual-drug controllable co-assembly nanosystem for targeted and synergistic treatment of hepatocellular carcinoma.

The effectiveness of chemotherapeutic agents for hepatocellular carcinoma (HCC) is unsatisfactory because of tumor heterogeneity, multidrug resistance, and poor target accumulation. Therefore, multimodality-treatment with accurate drug delivery has become increasingly popular. Herein, a cell penetrating peptide-aptamer dual modified-nanocomposite (USILA NPs) was successfully constructed by coating a cell penetrating peptide and aptamer onto the surface of sorafenib (Sora), ursolic acid (UA) and indocyanine green (ICG) condensed nanodrug (USI NPs) via one-pot assembly for targeted and synergistic HCC treatment. USILA NPs showed higher cellular uptake and cytotoxicity in HepG2 and H22 cells, with a high expression of epithelial cell adhesion molecule (EpCAM). Furthermore, these NPs caused more significant mitochondrial membrane potential reduction and cell apoptosis. These NPs could selectively accumulate at the tumor site of H22 tumor-bearing mice and were detected with the help of ICG fluorescence; moreover, they retarded tumor growth better than monotherapy. Thus, USILA NPs can realize the targeted delivery of dual drugs and the integration of diagnosis and treatment. Moreover, the effects were more significant after co-administration of iRGD peptide, a tumor-penetrating peptide with better penetration promoting ability or programmed cell death ligand 1 (PD-L1) antibody for the reversal of the immunosuppressive state in the tumor microenvironment. The tumor inhibition rates of USILA NPs + iRGD peptide or USILA NPs + PD-L1 antibody with good therapeutic safety were 72.38 % and 67.91 % compared with control, respectively. Overall, this composite nanosystem could act as a promising targeted tool and provide an effective intervention strategy for enhanced HCC synergistic treatment.

Demonstration of posturographic parameters of squat-stand activity in hemiparetic patients on a new multi-utility balance assessing and training system.

BACKGROUND: Quantitative evaluation of position control ability in stroke patients is needed. Here we report a demonstration of position control ability assessment and test-retest reliability during squat-stand activity on a new system in hemiparetic patients and controls. METHODS: Sixty-two healthy adults and thirty-four hemiparetics were enrolled. RESULTS: During partial weight support, the ICCs ranged from 0.77 to 0.91, which indicated a good reliability. During standard weight bearing and resistance, the ICCs varied from 0.64 to 0.86 and 0.54 to 0.84, respectively, indicating a fair reliability. Compared with the healthy adults, the stroke patients demonstrated poorer position control ability. CONCLUSIONS: The posturography of the squat-stand activity is a new and reliable measurement tool for position control. According to the methods proposed here, hemiparetics can be differentiated from healthy adults using the squat-stand activity. This activity will provide a new evaluation tool and therapy with visual feedback for the stroke patients. TRIAL REGISTRATION: Chinese clinical trial registry, ChiCTR-TRC-10000863.

[Effects of early application of Tuina treatment on quadriceps surface myoelectricity in patients after total knee arthroplasty: a randomized controlled trial].

BACKGROUND: Total knee replacement surgery is commonly used in end-stage diseases of the knee. It is important for improving surgical efficacy and patient satisfaction by promoting early rehabilitation of patients and improving knee function. OBJECTIVE: To observe the effects of early application of Tuina treatment on quadriceps surface electromyography (EMG) in patients with rheumatoid arthritis having undergone total knee arthroplasty. DESIGN, SETTING, PARTICIPANTS AND INTERVENTION: The study was performed at the Orthopedic Department of Huashan Hospital, Fudan University, and the Shanghai Guanghua Hospital of Integrated Traditional Chinese and Western Medicine from June 2010 to September 2011. A total of 66 patients with rheumatoid arthritis who had undergone total knee replacement surgery were randomly divided into control group and observation group, 33 cases in each. The patients in the control group were administered with continuous passive training (CPM), and the patients in the observation group were treated with CPM combined with Tuina, from prior surgery to four weeks post-surgery. MAIN OUTCOME MEASURES: The knee function was evaluated using the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) questionnaire at baseline and 4 weeks after the surgery. Quadriceps surface EMG was also detected at the same time points. RESULTS: After 4 weeks of Tuina and comprehensive rehabilitation intervention, the WOMAC questionnaire score of the observation group was decreased compared with the control group (P<0.01); median frequency and integrated electromyography of the rectus femoris and vastus medialis muscles, which were recorded by EMG, in the observation group were higher than those in the control group (P<0.01). CONCLUSION: Tuina can improve the recovery of patients who have undergone total knee replacement by increasing quadriceps EMG.

Treadmill pre-training suppresses the release of glutamate resulting from cerebral ischemia in rats.

This study was designed to investigate the neuroprotective effect of treadmill pre-training against the over-release of glutamate resulting from cerebral ischemia. Sprague-Dawley rats underwent 2 weeks of treadmill run-training before cerebral ischemia was performed by middle cerebral artery occlusion. The level of glutamate in brain extracellular fluid was detected before, during and after ischemia/reperfusion. The expression of metabotropic glutamate receptor-1 (mGluR1) mRNA in striatum was examined after ischemia for 80 min and reperfusion for 240 min. Neurological defect score and brain infarction volumes were measured. The treadmill pre-training significantly suppressed the release of glutamate, and reduced the expression of mGluR1 mRNA at 59% (P < 0.01) and 62% (P < 0.05), respectively, as compared with the ischemia group. The neurological defect score and infarction volume were significantly improved by 75% (P < 0.01) and 74% (P < 0.01), respectively, in the pre-training group, as compared to the ischemia group. Treadmill pre-training has a significant neuroprotective function against ischemia/reperfusion injury, by suppressing glutamate release resulting from cerebral ischemia, and this effect may be mediated by downregulation of mGluR1.

Pre-ischemic treadmill training induces tolerance to brain ischemia: involvement of glutamate and ERK1/2.

Physical exercise has been shown to be beneficial in stroke patients and animal stroke models. However, the exact mechanisms underlying this effect are not yet very clear. The present study investigated whether pre-ischemic treadmill training could induce brain ischemic tolerance (BIT) by inhibiting the excessive glutamate release and event-related kinase 1/2 (ERK1/2) activation observed in rats exposed to middle cerebral artery occlusion (MCAO). Sprague-Dawley rats were divided into three groups (n = 12/group): sham surgery without prior exercise, MCAO without prior exercise and MCAO following three weeks of exercise. Pre-MCAO exercise significantly reduced brain infarct size (103.1 +/- 6.7 mm3) relative to MCAO without prior exercise (175.9 +/- 13.5 mm3). Similarly, pre-MCAO exercise significantly reduced neurological defects (1.83 +/- 0.75) relative to MCAO without exercise (3.00 +/- 0.63). As expected, MCAO increased levels of phospho-ERK1/2 (69 +/- 5%) relative to sham surgery (40 +/- 5%), and phospho-ERK1/2 levels were normalized in rats exposed to pre-ischemic treadmill training (52 +/- 6%) relative to MCAO without exercise (69% +/- 5%). Parallel effects were observed on striatal glutamate overflow. This study suggests that pre-ischemic treadmill training might induce neuroprotection by inhibiting the phospho-ERK1/2 over-activation and reducing excessive glutamate release.

Protective effect of tetraethyl pyrazine against focal cerebral ischemia/reperfusion injury in rats: therapeutic time window and its mechanism.

INTRODUCTION: Tetramethyl pyrazine has been considered an effective agent in treating neurons ischemia/reperfusion injury, but the mechanism of its therapeutic effect remains unclear. This study was to explore the therapeutic time window and mechanism of tetramethyl pyrazine on temporary focal cerebral ischemia/reperfusion injury. MATERIALS AND METHODS: Middle cerebral artery occlusion was conducted in male Sprague-Dawley rats and 20 mg/kg of tetramethyl pyrazine was intraperitoneally injected at different time points. At 72 h after reperfusion, all animals' neurologic deficit scores were evaluated. Cerebrums were removed and cerebral infarction volume was measured. The expression of thioredoxin and thioredoxin reductase mRNA was determined at 6 and 24 h after reperfusion. RESULTS: Cerebral infarction volume and neurological deficit scores were significantly decreased in the group with tetramethyl pyrazine treatment. The expression of thioredoxin-1/thioredoxin-2 and thioredoxin reductase-1/thioredoxin reductase-2 was significantly decreased in rats with ischemia/reperfusion injury, while it was increased by tetramethyl pyrazine administration. CONCLUSIONS: Treatment with tetramethyl pyrazine, within 4 h after reperfusion, protects the brain from ischemic reperfusion injury in rats. The neuroprotective mechanism of tetramethyl pyrazine treatment is, in part, mediated through the upregulation of thioredoxin transcription.

[Clinical study of standardized tertiary rehabilitation program in promoting upper and lower limbs motor function in stroke patients].

OBJECTIVE: To investigate the effect of standardized tertiary rehabilitation (STR) on the upper and lower limbs' motor function in the patients with cerebral stroke accompanied by hemiplegia. METHODS: 1365 patients with cerebral stroke accompanied by hemiplegia enrolled sequentially from 22 hospitals were classified into two groups: cerebral infarction group and hemorrhage group, and then randomly subdivided into two groups: test group (n = 688, administered with STR in addition to routine interventions of neurological medicine) and control group (n = 677, given the routine interventions). All patients were assessed with the scale of Modified Fugl-Meyer Motor Assessment (M-FMMA) at the time of enrollment and 1, 3, and 6 months after the stroke respectively. The evaluation was blind on the part of the physician. RESULTS: At each of the following post-stroke checkpoints, the scores of the functions in the cerebral infarction or hemorrhage test group were higher than those in the control (P < 0.01). The M-FMMA scores at the enrollment, and 1, 3, and 6 months after stroke of the cerebral infarction patients in the test group were 28, 47, 65, and 75 respectively, and the M-FMMA scores at the enrollment, and 1, 3, and 6 months after stroke of the cerebral infarction patients in the control group were 26, 37, 48, and 55 respectively. The M-FMMA scores at the enrollment, and 1, 3, and 6 months after stroke of the cerebral hemorrhage patients in the test group were 23, 44, 67, and 80 respectively, and the M-FMMA scores at the enrollment, and 1, 3, and 6 months after stroke of the cerebral hemorrhage patients in the control group were 21, 32, 46, and 55 respectively. During the STR, the scores of functional improvement of both the cerebral infarction and hemorrhage test groups were higher than those of the control groups (both P < 0.01). By the end of the 6th month after stroke, the scores of functional improvement of both the cerebral infarction and hemorrhage test groups went up by 47 and 56 respectively for the 2 test groups, while went up by 29 and 34 respectively for the 2 control groups. Compared with the control groups, the score of functional improvement of the cerebral infarction test group was 18 points higher, and that of the cerebral hemorrhage test group was 22 points higher. CONCLUSION: STR significantly improves the neurological function in the patients with cerebral strokes accompanied by hemiplegia.

Constraint-induced movement therapy promotes motor function recovery and downregulates phosphorylated extracellular regulated protein kinase expression in ischemic brain tissue of rats.

Motor function impairment is a common outcome of stroke. Constraint-induced movement therapy (CIMT) involving intensive use of the impaired limb while restraining the unaffected limb is widely used to overcome the effects of 'learned non-use' and improve limb function after stroke. However, the underlying mechanism of CIMT remains unclear. In the present study, rats were randomly divided into a middle cerebral artery occlusion (model) group, a CIMT + model (CIMT) group, or a sham group. Restriction of the affected limb by plaster cast was performed in the CIMT and sham groups. Compared with the model group, CIMT significantly improved the forelimb functional performance in rats. By western blot assay, the expression of phosphorylated extracellular regulated protein kinase in the bilateral cortex and hippocampi of cerebral ischemic rats in the CIMT group was significantly lower than that in the model group, and was similar to sham group levels. These data suggest that functional recovery after CIMT may be related to decreased expression of phosphorylated extracellular regulated protein kinase in the bilateral cortex and hippocampi.

[Effects of standardized three-stage rehabilitation on recovery of neurological function in stroke patients with hemiplegia].

OBJECTIVE: To explore the effects of standardized three stages' rehabilitation on the neurological function in stroke patients with hemiplegia. METHODS: All 52 patients firstly are brought into two blocks: primary cerebral infarction and primary cerebral hemorrhage then are divided into treated group and controlled group randomly. Patients in the treated group are given Standardized Three Stages' Rehabilitation, while those in the controlled group are only given normal internal medicine treatments that are the same as the treated group but without Standardized Three Stages' Rehabilitation. All patients would be assessed with the scale of Clinical Neurological Function Defects (CNFD) at the entering time, the end of 1st month, 3rd month and 6th month respectively after stroke. RESULTS: The scores of the treated group are lower than those of the controlled group (P < 0.001) at every stage, the margins between every stage's scores in the treated group are greater than those in the controlled group (P < 0.001). The scores of the treated group's patients are about 51%, 34%, 18% and 8% of total scores at the entering time, the end of 1st month, 3rd month and 6th month after stroke differently, but that of the controlled group are about 58%, 54%, 42% and 37% of total scores differently. The margins between the scores of entering time and that of the end of 1st month, 3rd month and 6th month in the treated group are 17%, 33% and 43% of total scores respectively, but that of the controlled group are about 5%, 16% and 21% of total scores differently. CONCLUSION: Standardized three stages' rehabilitation could promote stroke patients' motor function of every stage obviously.

Long-term three-stage rehabilitation intervention alleviates spasticity of the elbows, fingers, and plantar flexors and improves activities of daily living in ischemic stroke patients: a randomized, controlled trial.

To investigate the effects of rehabilitation interventions on spasticity and activities of daily living (ADL) in ischemic stroke patients. A total of 165 ischemic stroke patients were recruited and assigned randomly to a control group (CG, n=82) or a therapeutic group (TG, n=83). Rehabilitation interventions were performed in the TG. The Modified Ashworth Scale was used to evaluate the severity of spasticity in the fingers, elbows, and plantar flexors, and the Modified Barthel Index (MBI) was used to measure ADL performance. Evaluations were performed at baseline (M0) and at the end of the first, third, and sixth months (M1, M3, M6) after enrollment. At M0, 20.8% (16/77) in the CG and 29.9% (23/77) in the TG developed spasticity, whereas at M6, the incidence of spasticity increased to 36.4% (28/77) in the TG and 42.9% (33/77) of patients in the CG. Fewer patients developed spasticity in the fingers, elbows, and ankles in the TG than CG, respectively. Both groups showed significant improvements in MBI scores (M6 vs. M0, P<0.01). MBI scores correlated negatively with the severity of spasticity in both groups at M6. Long-term standardized rehabilitation interventions alleviate spasticity and promote ADL with the presence of minor spasticity (Supplementary video, Supplemental digital content 1, http://links.lww.com/WNR/A291).

Prospective, randomized controlled trial of physiotherapy and acupuncture on motor function and daily activities in patients with ischemic stroke.

OBJECTIVE: To assess the value of acupuncture for promoting the recovery of patients with ischemic stroke and to determine whether the outcomes of combined physiotherapy and acupuncture are superior to those with physiotherapy alone. DESIGN: Prospective randomized controlled trial. SETTING: Department of Rehabilitation Medicine, Huashan Hospital, Fudan University, P. R. China. PARTICIPANTS: 120 inpatients and outpatients (84 men and 36 women). INTERVENTIONS: Acupuncture, physiotherapy, and physiotherapy combined with acupuncture. MAIN OUTCOME MEASURES: Motor function in the limbs was measured with the Fugl-Meyer assessment (FMA). The modified Barthel index (MBI) was used to rate activities of daily living. All evaluations were performed by assessors blinded to treatment group. RESULTS: On the first day of therapy (day 0, baseline), FMA and MBI scores did not significantly differ among the treatment groups. Compared with baseline, on the 28th day of therapy the mean FMA scores of the physiotherapy, acupuncture, and combined treatment groups had increased by 65.6%, 57.7%, and 67.2%, respectively; on the 56th day, FMA scores had increased by 88.1%, 64.5%, and 88.6%, respectively (p<0.05). The respective MBI scores in the three groups increased by 85.2%, 60.4%, and 63.4% at day 28 and by 108.0%, 71.2%, and 86.2% at day 56, respectively (p<0.05). However, FMA scores did not significantly differ among the three treatment groups on the 28th day. By the 56th day, the FMA and MBI scores of the physiotherapy group were 46.1% and 33.2% greater, respectively, than those in the acupuncture group p<0.05). No significant differences were seen between the combined treatment group and the other groups. In addition, the FMA subscores for the upper extremities did not reflect any significant improvement in any group on the 56th day. Although the FMA subscores for the upper and lower extremities and the MBI score in the combined treatment group were higher than those in the acupuncture group, the differences were not statistically significant. CONCLUSIONS: Acupuncture is less effective for the outcome measures studied than is physiotherapy. Moreover, the therapeutic effect of combining acupuncture with physiotherapy was not superior to that of physiotherapy alone. A larger-scale clinical trial is necessary to confirm these findings.

Pre-ischemic treadmill training affects glutamate and gamma aminobutyric acid levels in the striatal dialysate of a rat model of cerebral ischemia.

AIMS: Treadmill training has been shown to improve function in animal models and patients with cerebral ischemia. However, the neurochemical effects of this intervention on the ischemic brain have not been well studied. This study was designed to evaluate the effects of pre-ischemic treadmill training on the release of glutamate and gamma-aminobutyric acid (GABA) from the striatum in a rat middle cerebral artery occlusion (MCAO) model. MAIN METHODS: Rats were divided into five groups: sham control without MCAO, and 0, 1, 2 and 4 weeks pre-ischemic treadmill training. After training, cerebral ischemia was induced by MCAO for 120 min, followed by reperfusion. Microdialysis was used to collect dialysates from the striatum immediately before ischemia, and at 40, 80 and 120 min after ischemia, as well as at 40, 80, 120, 160, 200 and 240 min after reperfusion. KEY FINDINGS: Pre-ischemic treadmill training decreased glutamate release and increased GABA release during the acute phase of cerebral ischemia/reperfusion. Treadmill training for at least 2 weeks produced statistically significant changes in GABA/glutamate release. SIGNIFICANCE: The present study suggests that treadmill training inhibits the excessive release of glutamate, by stimulating GABA release during the acute phase of cerebral ischemia. This may be one of the important mechanisms to protect the striatal neurons from ischemic damage.