Ilizarov technology in China: a historic review of thirty-one years.

PURPOSE: To summarize the evolution of Ilizarov technology in China, highlight important milestones, introduce the atmosphere of the era concerning the first uses and development of this technology, and share Chinese modification and experience in this field. METHOD: A thorough interview with senior ASAMI members of China and literature search and physical books in libraries was undertaken to summarize the history of Ilizarov technology in China. RESULTS: The formal development of Ilizarov technology began when professor Ilizarov himself came to Beijing (1991) and gave a speech. In the following 31 years, his technology was rapidly developed through China, with many symposiums held and associations established including ASAMI China (2003) and ILLRS China (2015). Today, Ilizarov technology has become the main treatment of complex fractures, defects, nonunion, infections, deformities, and chronic ischemic ulcers of the limbs. In those years, Chinese scholars also developed some special treatment methods and made many modifications to Ilizarov external fixators. CONCLUSION: Ilizarov technology has developed in China for 31 years. It revolutionized the treatment of complex limb traumas, deformities, and diseases. In the treatment of millions of patients, Chinese scholars had many unique experiences and made modifications to this technology which is worthy to share with the world.

Stat3-siRNA inhibits the growth of gastric cancer in vitro and in vivo.

Gastric cancer remains one of the most prevalent and lethal malignancies in the world. Despite new advances in treatment and diagnosis, patients with advanced gastric cancer are still difficult to cure resulting in a high mortality rate and poor prognosis. Signal transducer and activator of transcription 3 (Stat3) is observed aberrant in multiple tumours, including gastric cancer. Stat3 overexpression was confirmed performing a vital role in tumorigenesis. In the present study, we constructed a pSi-Stat3 plasmid to silence Stat3 and investigated the effect of pSi-Stat3 on cell proliferation, apoptosis and cell cycle progression in gastric cancer cell line SGC-7901 and mice xenograft model. Downstream proteins of Stat3, including Cyclin-D1, Survivin and Bcl-2, were detected as well for the underlying mechanism exploration. It showed that pSi-Stat3 can effectively silence the expression of Stat3 and inhibits the growth of gastric tumour both in vitro and in vivo significantly via cell apoptosis and cell cycle shift induction. The findings suggest that Stat3 signal pathway might be a promising therapeutic target for tumour treatment, including gastric cancer.

[Effects of platycodin D in combination with different active ingredients of Chinese herbs on proliferation and invasion of 4T1 and MDA-MB-231 breast cancer cell lines].

OBJECTIVE: To investigate the effects of platycodin D in combination with different active ingredients of Chinese herbs under different therapeutic principles on proliferation and invasion of 4T1 and MDA-MB-231 breast cancer cell lines. METHODS: The effective doses of platycodin D, Ophiopogon total saponins, curcumenol and osthole in inhibiting proliferation of breast cancer cell lines 4T1 and MDA-MB-231 were detected by methyl thiazolyl tetrazolium (MTT) assay, respectively. Optimized combinations of platycodin D with Ophiopogon total saponins, curcumenol, or osthole were determined by uniform design method. Effects of the optimized combinations of platycodin D with the three ingredients on proliferation and invasion of 4T1 and MDA-MB-231 cells were verified and evaluated by MTT assay and Transwell chamber test, respectively. RESULTS: Verifying study showed that the inhibitory effects of platycodin D in combination with curcumenol or osthole on proliferation of 4T1 and MDA-MB-231 cells were better than those of platycodin D in combination with Ophiopogon total saponins and each ingredient used alone (P<0.05 or P<0.01). The inhibitory effect of platycodin D in combination with Ophiopogon total saponins or osthole on invasion of 4T1 cells was significantly better than those of platycodin D in combination with curcumenol and each ingredient used alone (P<0.05 or P<0.01). Moreover, the inhibitory effect of platycodin D in combination with curcumenol or osthole on invasion of MDA-MB-231 cells was significantly better than that of platycodin D in combination with Ophiopogon total saponins (P<0.01). CONCLUSION: The optimized combinations of platycodin D with three different active ingredients of Chinese herbs under different therapeutic principles can significantly inhibit the proliferation and decrease the invasion of 4T1 and MDA-MB-231 cells. Different platycodin D combinations have different potency in suppressing breast cancer cell proliferation and invasion.

[Inhibitory effects of osthole, psoralen and aconitine on invasive activities of breast cancer MDA-MB-231BO cell line and the mechanisms].

OBJECTIVE: To explore the inhibitory effects and to investigate the mechanisms of combined treatment of osthole, psoralen with aconitine on human breast cancer cell line MDA-MB-231BO. METHODS: The best inhibitory concentration of osthole, psoralen combined with aconitine on MDA-MB-231BO cells was obtained by stepwise regression analysis after adopting a uniform experiment design. The invasive activities were observed by transwell assays, and expressions of transforming growth factor-ß1 (TGF-ß1), Smad2, Smad3, Smad4, Smad7, nuclear factor-κB (NF-κB) and receptor activator of NF-κB ligand (RANK) mRNAs were analyzed by real-time quantitative polymerase chain reaction. RESULTS: The optimal combination concentrations of osthol, psoralen and aconitine were 6.44, 8.89 and 9.44 µg/mL, respectively. Cell invasion was significantly inhibited after 24 hours of treatment using the combination drugs and zoledronic acid. TGF-ß1, Smad2, Smad3, Smad4, Smad7, NF-κB and RANK mRNA expressions of the optimal combination group and zoledronic acid group were significantly reduced compared with the control group (P<0.01). Furthermore, TGF-ß1, Smad2, Smad3, Smad4, Smad7, NF-κB and RANK mRNA expressions of the optimal combination group were significantly lower than those of the weak combination group (P<0.01). CONCLUSION: Combination treatment of osthole, psoralen with aconitine can inhibit cancer cell invasion, which is a result of alteration of the TGF-ß/Smad signaling pathway and down-regulation of NF-κB and RANK expressions.

[Clinical analysis of 107 patients with foot and ankle deformities caused by spinal bifida].

OBJECTIVE: To analyze the incidence, clinical features, deformity categories and orthopedic treatment of foot and ankle deformities caused by spinal bifida. METHODS: The charts of the patients received surgical treatment between January 1990 and July 2009 were studied retrospectively, and the data were analyzed. RESULTS: One hundred and seven cases of foot and ankle deformities caused by spinal bifida received surgical treatment and were included. There were 44 male and 63 female patients. The average age was 17.7 years (range, 1.3 - 52.0 years). And 50.5% (54/107) of cases were over 18 years old and had spinal bifida occulta, and the other 49.5% had spinal bifida manifesta. There was only one case of thoracic spinal bifida (T(3-8)), while the other 106 cases had lumbosacral vertebrae cleft (mainly L(3) to Sacrum). Among a total of 165 feet, unilateral involvement was found in 49 cases (22 cases on the left side, 27 cases on the right side), bilateral involvement in 58 cases. Combined ankle-foot deformities included 76 varus talipes, 23 talipes valgus, 15 flail feet, and 51 other foot deformities. Other site deformities, as a result of spinal bifida, included knee flexion or hyperextension deformity in 4 cases, hip deformity (hip adduction, flexion, or hip dislocation, pelvic tilt, lower extremity discrepancy, etc.) in 17 cases, and urinatory dysfunction and defecation in 30 cases. Twenty-nine of 54 cases with spinal bifida occulta failed to be diagnosed in other hospitals and the misdiagnosed rate reached 53.7% (29/54). Corrective surgery was performed in only 26 patients. And 50.5% (54/107) of patients (over 18 years old) had severe foot and ankle deformities due to a failure of prior surgical treatment. CONCLUSIONS: Spinal bifida is the most commonly found in the lumbosacral vertebrae. Although the main pathogenesis is developmental abnormalities of spinal cord and nerve, the secondary deformity is usually located on the foot and ankle. Some young orthopedic surgeons may not have enough awareness and treatment experience about this disease due to over-specialty of the orthopaedics, so the delay of early diagnosis and treatment is often found and many severe foot and ankle deformities occur.

[Analysis of middle-term outcome of the imaging characteristics after the percutaneous laser disc decompression].

OBJECTIVE: To study the imaging findings outcome of the percutaneous laser disc decompression (PLDD) and evaluate the middle-term safety and efficacy of PLDD. METHODS: The imaging data of 22 cases suffered cervical spondylosis or lumbar spondylosis and treated by PLDD more than 3 years were retrospectively reviewed. To observe the intervertebral space of the invasive disc and the intervertebral disc hernia before and after the operation on the lateral projection of X-ray and the axial view of the MRI. To make a statistical analysis of the data. RESULTS: There was no obvious change of disc height involving the anterior disc height, the intermediate disc height and the posterior disc height after the PLDD at the final follow-up (P > 0.05). At last time follow-up, the Sagittal Index (SI) are 0.10 - 0.54 and 0.06 - 0.39 before and after the PLDD in the treatment of cervical vertebra. The statistical difference were significant (P < 0.05). In lumbar vertebra, the SI is 0 - 0.71 and 0 - 0.48 disc before and after the PLDD. The statistical difference were very significant (P < 0.01). CONCLUSIONS: The PLDD will not destroy the intervertebral space of the cervical and lumbar vertebra obviously; moreover, it can effectively reduce the disk hernia. The PLDD is a safe and effective Mini-invasive surgery for cervical vertebra and lumbar vertebra diseases.

Application of neural stem cells in tissue-engineered artificial nerve.

OBJECTIVE: To observe the curative effect of neural stem cells (NSCs), which are used in tissue-engineered artificial nerve, on repairing rabbit 10-mm facial nerve defects. METHODS: Thirty-six Oryctolagus cuniculi were randomly divided into three groups (each group with 12 Oryctolagus cuniculi). In group A, chitosan conduit, collagen protein sponge, nerve growth factor (NGF), and NSCs were used. In group B, chitosan conduit, collagen sponge, and NGF were used. In group C, nerve autograft was performed. Electrophysiologic detection, histologic observation, and BrdU and S100 immunohistochemical examination were performed 12 weeks after operation. RESULTS: All observation items in group A were better than those in group B (P < 0.01), and there were no significant differences between group A and group C (P > 0.05). CONCLUSION: NSCs may be served as seed cells of peripheral nerve tissue engineering and be used in artificial nerve to repair facial nerve defects.

[Treatment of equinovarus accompanied with ulcer using Ilizarov technology and limited operation].

OBJECTIVE: To explore the surgical method and clinical curative effect of Ilizarov technique combined with limited surgical treatment for neurotrophic malnutrition equinovarus accompanied with weight-bearing area ulcer. METHODS: From July 2004 to December 2011, 21 cases of neurological disorders equinovarus with skin ulcer in weight-bearing area were treated including 14 males and 7 females with an average age of 21.3 years old ranging from 8 to 32 years. Among them,19 cases with talipes equinovarus were on account of spina bifid and 2 cases on account of spinal cord injury of lumbar sacral segment. Nine cases were on the left foot and 12 cases were right foot. The ulcer area in out-below side of the fifth metatarsal bone was in 6 cases, in basement of the fifth metatarsal bone in 5 cases,in lateral of calcaneocuboid joint 2 cases, and in lateral of subtalar joint in 8 cases. Surgical procedure included plantar medial release technique, tendo-chilles lengthenning, and three-joint partial osteotomy. During operation,the skin and soft tissue of ulcer were resected and the incision was sutured when the ulcer was in the lateral of subtalar joint, and ankle Ilizarov external fixation for extension was installed after correcting talipes equinovarus partially. For the legacy skin ulcer in weight-bearing area,the Shenrg-ji cream (Chinese characters) was used after operation for one time per day until the wound healed. Adjusting Ilizarov external fixation for correcting residual deformity until it is satisfaction. During this process weight-bearing walking was keeping. RESULTS: All cases were followed up from 6 to 52 months with an average of 28 months. All ulcers were healing and no recurrence. The ulcer healing time was 14 to 36 days postoperative (26 days in average). Eighteen deformity feet were corrected satisfactorily, and recovered fully plantar foot, 3 feet appeared mild deformity after 1 year. Ankle joint activity appeared limited in 15 cases and got completely rigidity in 6 cases. The AOFAS increased from preoperative 34.0±7.2 to 86.0±8.5 postoperatively; 8 feet got excellent, 10 good,3 fair. CONCLUSION: The clinical effect of Ilizarov technique combined with limited operation and the Sheng-ji cream in correction of the talipes equinovarus with skin ulcer in weigh-bearing area on account of neurotrophic malnutrition is satisfactory, the surgical method is simple and the treating course in security, and serious complications can be avoided.

Apoptotic effects of psiRNA-STAT3 on 4T1 breast cancer cells in vitro.

BACKGROUND: The aim of this study was to investigate the effect of a Lipofectamine2000 (Life2000) Transfection Reagent transfected psiRNA-STAT3 plasmid on 4T1 breast cancer cells. MATERIALS AND METHODS: MTT was used to detect the cell proliferation of breast cancer 4T1 cells at different periods (0h, 6h, 8h, 10h); the cell cycle was assessed by flow cytometry; variation of apoptosis and mitochondrial membrane potential was observed under a fluorescence microscope; immunohistochemical staining was used to determine the expression of caspase-3 and cyclin-D1 protein. RESULTS: An obvious effect of inhibition to 4T1 cancer cells could be observed at 8h after the psiRNA-STAT3 was transfected. Typical alterations of apoptotic morphological features were visible in the psiRNA-STAT3 treatment group. Mitochondrial membrane potential decreased significantly, the number of cells was increased in G0/G1 phase, and the number of cells was decreased in S phase, and the data were statistically significant (p<0.05), compared with the Scramble and Mock groups. Expression of caspase-3 protein was increased significantly, while that of cyclin D1 was significantly decreased. CONCLUSIONS: Life2000 transfected psiRNA-STAT3 plasmid can inhibit 4T1 tumor cell proliferation and promote apoptosis of 4T1 tumor cells, which process depends on the regulation of expression of cyclin D1 and caspase-3 protein.

Novel esophageal squamous cell carcinoma bone metastatic clone isolated by scintigraphy, X ray and micro PET/CT.

AIM: To establish a Chinese esophageal squamous cell carcinoma (ESCC) cell line with high bone metastasis potency using (99m)Tc-methylene diphosphonate ((99m)Tc-MDP) micro-pinhole scintigraphy, X ray and micro-positron emission tomography/computed tomography (PET/CT) for exploring the mechanism of occurrence and development in esophageal cancer. METHODS: The cells came from a BALB/c nu/nu immunodeficient mouse, and oncogenic tumor tissue was from a surgical specimen from a 61-year-old male patient with ESCC. The cell growth curve was mapped and analysis of chromosome karyotype was performed. Approximately 1 × 106 oncogenic cells were injected into the left cardiac ventricle of immunodeficient mice. The bone metastatic lesions of tumor-bearing mice were detected by (99m)Tc-MDP scintigraphy, micro-PET/CT and X-ray, and were resected from the mice under deep anesthesia. The bone metastatic cells in the lesions were used for culture and for repeated intracardiac inoculation. This in vivo/in vitro experimental metastasis study was repeated for four cycles. All of the suspicious bone sites were confirmed by pathology. Real-time polymerase chain reaction was used to compare the gene expression in the parental cells and in the bone metastatic clone. RESULTS: The surgical specimen was implanted subcutaneously in immunodeficient mice and the tumorigenesis rate was 100%. First-passage oncogenic cells were named CEK-Sq-1. The chromosome karyotype analysis of the cell line was hypotriploid. The bone metastasis rate went from 20% with the first-passage oncogenic cells via intracardiac inoculation to 90% after four cycles. The established bone metastasis clone named CEK-Sq-1BM had a high potential to metastasize in bone, including mandible, humerus, thoracic and lumbar vertebrae, scapula and femur. The bone metastasis lesions were successfully detected by micro-pinhole bone scintigraphy, micro-PET/CT, and X-ray. The sensitivity, specificity and accuracy of the micro-pinhole scintigraphy, X-ray, and micro-PET/CT imaging examinations were: 89.66%/32%/80%, 88.2%/100%/89.2%, and 88.75%/77.5%/87.5%, respectively. Some gene expression difference was found between parental and bone metastasis cells. CONCLUSION: This newly established Chinese ESCC cell line and animal model may provide a useful tool for the study of the pathogenesis and development of esophageal carcinoma.

Plasmid-based Survivin shRNA and GRIM-19 carried by attenuated Salmonella suppresses tumor cell growth.

Persistent activation of Survivin and its overexpression contribute to the formation, progression and metastasis of several different tumor types. Therefore, Survivin is an ideal target for RNA interference mediated-growth inhibition. Blockade of Survivin using specific short hairpin RNAs (shRNA) can significantly reduce prostate tumor growth. RNA interference does not fully ablate target gene expression, owing to the idiosyncrasies associated with shRNAs and their targets. To enhance the therapeutic efficacy of Survivin-specific shRNA, we employed a combinatorial expression of Survivin-specific shRNA and gene associated with retinoid-interferon-induced mortality-19 (GRIM-19). Then, the GRIM-19 coding sequences and Survivin-specific shRNAs were used to create a dual expression plasmid vector and were carried by an attenuated strain of Salmonella enteric serovar typhimurium (S. typhimurium) to treat prostate cancer in vitro and in vivo. We found that the co-expressed Survivin-specific shRNA and GRIM-19 synergistically and more effectively inhibited prostate tumor proliferation and survival, when compared with treatment with either single agent alone in vitro and in vivo. This study has provided a novel cancer gene therapeutic approach for prostate cancer.

Is XMRV a causal virus for prostate cancer?

The potential association between xenotropic murine leukaemia virus-related gammaretrovirus (XMRV) and prostate cancer (PCa) has been documented since 2006. It is important for furthering our understanding of the biological mechanisms of PCa to ascertain whether this association is causal. To summarize the available information on the epidemiological and laboratory findings of the association, we conducted a literature search of the PubMed electronic database (from March 2006 to February 2011) to identify relevant published studies that examined the association between XMRV and PCa. Although several studies showed the positive association between XMRV and PCa, more recent studies did not support this conclusion. The positive findings might be due to contamination of human samples. Further studies are needed to clarify this association.

Tacrine and bis(7)-tacrine attenuate cycloheximide-induced amnesia in mice, with attention to acute toxicity.

Effects of tacrine and bis(7)-tacrine (0.25-20 µmol/kg, s.c.) on cognitive behaviour in cycloheximide (CYH)-treated mice were investigated. Cognitive behaviour was assessed by open-field test and step-through task with a 24-hr retention interval. Drugs or vehicle was given 30 min. prior to the first session. Although CYH treatment (110 mg/kg, i.p.) alone did not affect the locomotor activity of mice, CYH treatment in combination with tacrine (20 µmol/kg) decreased the locomotor activity by 37% in the acquisition session, when compared with mice treated with CYH alone. Bis-(7) tacrine cotreatment did not produce any detectable effect on locomotor activity. During the retention trial, tacrine (5 µmol/kg) or bis(7)-tacrine (1 µmol/kg) enhanced the retention latency (by 3.8- or 1.4-fold, respectively) in CYH-treated mice. In both training and retention trials, CYH treatment increased the number of footshocks (by 50% and 11.3-fold, respectively). However, during the retention (but not training) trial, tacrine (5 µmol/kg) or bis(7)-tacrine (1 µmol/kg) decreased the footshocks (by 8.6-fold or 39%, respectively) in CYH-treated mice. Combined treatment with CYH and bis(7)-tacrine (but not tacrine) resulted in an increased mortality rate in mice. The results indicated that tacrine and bis(7)-tacrine improved the amnesia caused by CYH treatment. However, the combined treatment with bis(7)-tacrine and CYH administration caused acute toxicity.

Effective kinetics of schisandrin B on serum/hepatic triglyceride and total cholesterol levels in mice with and without the influence of fenofibrate.

Schisandrin B, an active ingredient isolated from the fruit of Schisandra chinensis, increased serum and hepatic triglyceride levels in mice. In the present study, the effective kinetics of schisandrin B on serum/hepatic triglyceride and total cholesterol levels in mice without and with the influence of fenofibrate were investigated. Parameters on hepatic index (the ratio of liver weight to body weight × 100) were also analyzed. Mice were intragastrically treated with schisandrin B at a single dose of 0.2, 0.4, 0.8, or 1.6 g/kg, without or with fenofibrate pretreatment (0.1 g/kg/day for 4 days, p.o.). Twenty-four hours after schisandrin B treatment, serum/hepatic triglyceride and total cholesterol levels were measured. Schisandrin B treatment dose-dependently increased serum and hepatic triglyceride levels as well as hepatic index in mice. In contrast, hepatic total cholesterol levels were decreased in a dose-dependent manner in schisandrin B-treated mice. Data obtained from effective kinetics analysis indicated that the action of schisandrin B on serum triglyceride had a higher specificity than those on hepatic total cholesterol and hepatic index. While fenofibrate pretreatment inhibited the schisandrin B-induced elevation in serum triglyceride levels, it completely abrogated the elevation of hepatic triglyceride levels in schisandrin B-treated mice. The combined treatment with schisandrin B and fenofibrate decreased hepatic total cholesterol level and increased the hepatic index in an additive or semi-additive manner, respectively. In conclusion, the results of effective kinetics analysis indicated that the schisandrin B-induced hypertriglyceridemia was competitively inhibited by fenofibrate. Schisandrin B may offer the prospect of setting up a mouse model of hypertriglyceridemia and fatty liver for screening triglyceride-lowering drug candidates.

Tacrine treatment at high dose suppresses the recognition memory in juvenile and adult mice with attention to hepatotoxicity.

It is well established that cholinergic over-stimulation can interfere with memory processes. The aim of this study was to evaluate the effect of tacrine, an acetylcholinesterase inhibitor, on recognition memory as well as the associated hepatotoxicity in juvenile (20-day-old) and adult (100-day-old) ICR male mice. Recognition memory was assessed by open-field test and step-through task without footshocks for three sessions between 08:00 and 13:00, with a 24-hr retention interval. Tacrine (10 or 40 µmol/kg) or vehicle was administered (s.c.) 20 min. prior to the first session. During the acquisition session, tacrine suppressed the open-field behaviours, including locomotor activity, rearing, grooming and defecation (by 77-100%) in mice of both ages. During the recall (observable in both ages) and re-recall (observable in juvenile mice) session, the locomotor activity and rearing number were significantly increased, indicative of impairment in recognition memory, in mice treated with tacrine 40 µmol/kg. During the training trial, tacrine decreased the step-through number in mice of both ages. In contrast, during the retention and re-retention trials, the step-through number was increased (by 92% and 93%, respectively), indicative of impairment in step-through memory, in juvenile but not adult mice treated with tacrine 40 µmol/kg. Tacrine 40 µmol/kg elevated the serum alanine aminotransferase (ALT) activity (by 135%) in juvenile mice, but reduced the ALT activity (by 42%) in adult mice. The results indicated that 20-day-old mice seemed to be more sensitive than 100-day-old mice to tacrine-induced impairment in recognition memory and the associated liver damage.

Plasmid-based Stat3 siRNA delivered by hydroxyapatite nanoparticles suppresses mouse prostate tumour growth in vivo.

DNA vector-based Stat3-specific RNA interference (si-Stat3) blocks Stat3 signalling and inhibits prostate tumour growth. However, the antitumour activity depends on the efficient delivery of si-Stat3. The effects on the growth of mouse prostate cancer cells of si-Stat3 delivered by hydroxyapatite were determined in this study. RM-1 tumour blocks were transplanted into C57BL/6 mice. CaCl2-modified hydroxyapatite carrying si-Stat3 plasmids were injected into tumours, and tumour growth and histology were determined. The expression levels of Stat3, pTyr-Stat3, Bcl-2, Bax, Caspase3, VEGF and cyclin D1 were measured by western blot analysis. Amounts of apoptosis in cancer cells were analysed with immunohistochemistry and the terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL) assay. The results showed that hydroxyapatite-delivered si-Stat3 significantly suppressed tumour growth up to 74% (P < 0.01). Stat3 expression was dramatically downregulated in the tumours. The immunohistochemistry and TUNEL results showed that si-Stat3-induced apoptosis (up to 42%, P < 0.01). The Stat3 downstream genes Bcl-2, VEGF and cyclin D1 were also strongly downregulated in the tumour tissues that also displayed significant increases in Bax expression and Caspase3 activity. These results suggest that hydroxyapatite can be used for the in vivo delivery of plasmid-based siRNAs into tumours.