The osteogenic niche-targeted arsenic nanoparticles prevent colonization of disseminated breast tumor cells in the bone

Up to 70% of patients with late-stage breast cancer have bone metastasis. Current treatment regimens for breast cancer bone metastasis are palliative with no therapeutic cure. Disseminated tumor cells (DTCs) colonize inside the osteogenic niches in the early stage of bone metastasis. Drug delivery into osteogenic niches to inhibit DTC colonization can prevent bone metastasis from entering its late stage and therefore cure bone metastasis. Here, we constructed a 50% DSS6 peptide conjugated nanoparticle to target the osteogenic niche. The osteogenic niche was always located at the endosteum with immature hydroxyapatite. Arsenic-manganese nanocrystals (around 14 nm) were loaded in osteogenic niche-targeted PEG-PLGA nanoparticles with an acidic environment-triggered arsenic release. Arsenic formulations greatly reduced 4T1 cell adhesion to mesenchymal stem cells (MSCs)/preosteoblasts (pre-OBs) and osteogenic differentiation of osteoblastic cells. Arsenic formulations also prevented tumor cell colonization and dormancy via altering the direct interaction between 4T1 cells and MSCs/pre-OBs. The chemotactic migration of 4T1 cells toward osteogenic cells was blocked by arsenic in mimic 3D osteogenic niche. Systemic administration of osteogenic niche-targeted arsenic nanoparticles significantly extended the survival of mice with 4T1 syngeneic bone metastasis. Our findings provide an effective approach for osteogenic niche-specific drug delivery and suggest that bone metastasis can be effectively inhibited by blockage of tumor cell colonization in the bone microenvironment.

The correlation between SUV measured on 18F-FDG PET/CT scan and residual tumor after argon-helium knife cryoablation / 介入放射学杂志

Objective To evaluate the correlation between standardized uptake value (SUV) measured on 18F-FDG PET/CT scan and residual tumor after argon-helium knife cryoablation.Methods The clinical data of 40 patients with hepatic or pulmonary malignant tumors,who were treated with argon-helium knife cryoablation during the period from March 2008 to December 2015 at authors' hospital,were collected.18FFDG PET/CT scan was performed both before and after the treatment,and the SUV values of each patient were calculated.The data were analyzed based on the pathological findings and clinical follow-up results.Results A total of 42 lesions were detected in the 40 patients.After treatment,radionuclide concentration was demonstrated in 38 lesions,and 16 lesions were proved to be the tumor residual by follow-up imaging and pathological examination.In other 22 lesions the radionuclide concentration was due to inflammatory response.The SUV value of the tumor residual was strikingly higher than that of the inflammatory response (6.13±1.21 vs.2.64±0.96,P＜0.05).The group with low SUV value had a lower recurrence rate (P=0.020) and a higher survival rate (P=0.039).The tumor survival rate of the low SUV value group was significantly lower than that of the group with high SUV value (x2=14.994,P=0.000 2).Conclusion 18F-FDG PET/CT imaging has unique value in promptly detecting marginal residual lesion after argon-helium knife cryoablation,which provides useful information for the evaluation of cryoablation effect as well as for the making of further therapeutic plan.

The effect of Connexin43 downregulation on biological functions of HUVEC / 药学学报

Connexin43 has been shown to play a pivotal role in wound healing process. Wound repair is enhanced by acute downregulation of connexin43, by increasing proliferation and migration of keratinocyte and fibroblast. Angiogenesis is also a central feature of wound repair, but little is known about the effects of connexin43 modulation on functions of endothelial cells. We used connexin43 specific small interference RNA (siRNA) to reduce the expression of connexin43 in human umbilical vein endothelial cell (HUVEC), and investigated the effects of connexin43 downregulation on intercellular communication, viability, proliferation, migration and angiogenic activity of HUVEC. Treatment of siRNA markedly reduced the expression of connexin43 by -80% in HUVEC (P < 0.05), and decreased the intercellular communication by -65% (P < 0.05). The viability, proliferation, migration and angiogenic activity of HUVEC decreased significantly (P < 0.05), compared with that of the normal cells. The results suggest that temporally downregulation of connexin43 expression at early stage of wound to inhibit the abnormal angiogenesis characterized with leaky and inflamed blood vessels, maybe a prerequisite for coordinated normal healing process.

Apolipoprotein M expression in human colorectal cancer tissues and its clinicopathological relevance / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the mRNA and protein expression levels of apolipoprotein M (apoM) in the human colorectal cancer tissues, and to explore its clinical relevance.</p><p><b>METHODS</b>Real-time PCR was carried out to determine the mRNA expression levels both in cancer tissue and its adjacent normal tissue from 20 patients with colorectal cancer. Immunohistochemistry was also carried out to determine the protein levels in 23 colorectal biopsy samples (7 normal mucosa, 6 inflammatory mucosa and 10 polyp tissues) and 20 cases of colorectal cancer tissues as well as the adjacent normal tissues.</p><p><b>RESULTS</b>Real-time PCR result showed that apoM mRNA level in the colorectal cancer tissues was significantly lower than that in their adjacent normal tissues (0.05±0.01 vs. 0.19±0.05, P<0.05). ApoM mRNA level in colorectal cancer tissues was statistically significant higher in the patients with lymph node metastasis as compared to the patients without lymph node metastasis (P<0.01). The median value of apoM protein in cancer tissues was 5.50, which was significantly lower than that in the adjacent normal tissues (10.5, P<0.05), inflammatory mucosa tissues (9.75, P<0.05), polyp tissues (11.0, P<0.01) and normal mucosa (10.5, P<0.05). No significant association was observed between the apoM protein level and the clinicopathological parameters of patients.</p><p><b>CONCLUSIONS</b>Both apoM mRNA and protein expression levels in colorectal cancer tissues are significantly decreased in contrast to normal and benign colorectal tissues. The apoM mRNA expression in colorectal cancer tissues is closely associated with nodal metastasis.</p>

Efficacy and safety of pancreas freezing / 中华胰腺病杂志

Objective To observe the blood biochemical and histological changes before and after pancreas freezing, to provide evidence for cryosurgery for pancreatic cancer. Methods Fifteen healthy pigs were divided into deep frozen group (n = 5), shallow frozen group (n = 5), non-frozen group (n = 3) and normal group (n = 2). After anesthesia and Iaparotomy, a probe of the Argon-Helium Surgical System was inserted into the pancreas, 100% and 10% argon output power were used in deep and shallow frozen group, respectively;and the temperature were - 130 ～ - 140℃ and - 110 ～ - 120℃, respectively;which results in an ice-ball with 15 ～ 20 mm in diameter. Then helium gas was inputted to increase the temperature to 10 ～ 20℃ for three minutes;then the whole process was repeated. A probe was inserted into the pancreas in the non-frozen group only and only laparotomy was performed in non-grozen group normal group and normal group. Serum amylase, IL-6, CRP levels before and after the experiment was determined;the pigs were sacrificed at day 7 and the pancreas was harvested for light microscope and electron microscope examination. Results The frozen pancreatic tissue became pitchy necrosis zone, and it could be distinguished from non-frozen tissue;there were obvious tissue necrosis in the center and para-center of frozen area, and the ultra-structure were destroyed and disappeared, mitochondria degranulation and rough endoplasmic reticulum degrannlation were observed. Serum amylase was elevated in 13 (86.7%) pigs and most returned to normal at 6th day. Serum IL-6 was slightly elevated in 5 (33.3%) pigs. There was no significant difference among all the groups in term of serum CRP. All the pigs were alive until the time of sacrifice. Conclusions Cryosurgery has affirmative fatal ablative effects on pancreatic tissue, and it is safe with no serious complications.

Treatment of hepatocellular carcinoma with transarterial chemoembolization and percutaneous cryosurgery sequential therapy / 中华消化杂志

Objective To evaluate the efficacy of transarterial chemoembolization (TACE) and percutaneous cryosurgery sequential therapy for unresectable hepatocellular carcinoma (HCC).Methods Four hundred and twenty patients with unresectable HCC were divided into sequential TACE-cryosurgery sequential (sequential) group (n=290) and cryosurgery alone (cryoalone) group (n = 130). TACE was performed with the routine operation; the percutaneous cryosurgery was conducted 2 to 4 weeks after TACE. The patients were followed up at the first month and once every 2 to 3 month later. Liver ultrasound or both computer tomography and alpha fetal protein were examined during follow-up. Results During a mean follow-up of (42±17) months (range from 24 to 70 months), the local recurrence rate of ablated lesion was 17% for all the patients, 11% and 24% for patients in sequential group and cryoalone groups respectively (P=0. 001). The overall 1-, 2-, 3-, 4-and 5-year survival rate was 72%, 57%, 47%, 39% and 31%, respectively. The 1- and 2-year survival rates (71% and 61 % ) in sequential group were similar to those (73 % and 54 % ) in cryo-alone group (P=0.69 and 0. 147), while the 4- and 5-year survival rates were higher in sequential group (49 % and 39 % ) than those (29 % and 23 % ) in cryo-alone group (P= 0.001). Eighteen patients with large HCC (＞5 cm in diameter) in sequential group survived for more than 5 years while no one in cryo-alone group. Complication rate was 24% in all patients, 21% and 26% for the sequential and cryo-alone groups respectively (P=0. 06). The incidence of hepatic bleeding was higher in cryo-alone group than in sequential group (P=0. 02). Liver crack occurred in two patients of the cryoalone group. Conclusions Pre-cryosurgical TACE increased the cryoablation efficacy and decrease its complications, especially hepatic bleeding. TACE and cryosurgery sequential therapy may be a better treatment for unresectable HCC, especially for large HCC.

Enhanced thymidine kinase gene vector and its killing effect on nasopharyngeal carcinoma in vitro and in vivo / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To construct a modified and enhanced thymidine kinase (TK) vector regulated by human telomerase catalytic subunit promoter (hTERT) promoter and cytomegalovirus (CMV) enhancer and its killing effect on nasopharyngeal carcinoma in vitro and in vivo and its safety in vivo.</p><p><b>METHODS</b>The pGL3-basic, as basic vector template, was linked and constructed into TK vector regulated by hTERT promoter and CMV enhancer with mono-promoter vector as control. Enhanced TK expression was confirmed by fluorescent microscopy and real time fluorescent quantitative PCR. Telomerase activity was measured by stretch PCR. Tumour killing effects were examined by MTT and Boyden areola. The effects of enhanced TK on the invasiveness of tumor cell NPC 5-8F and the growth of xenograft implanted in nude mice were investigated.</p><p><b>RESULTS</b>Compared with non-enhanced vector, TK expressed by the enhanced vector significantly increased in NPC 5-8F and MCF-7 cells, telomerase activity was positive in human in NPC 5-8F cells and breast cancer MCF-7 cells and negative in control human blood vessel endothelium ECV-304 cells. After ganciclovir(GCV) treatment, NPC 5-8F cell survival rate and invasiveness decreased and tumor progress of NPC xenograft implanted in nude mice was inhibited, without obvious toxicity effects on mouse liver and kidney.</p><p><b>CONCLUSIONS</b>The enhanced TK vector regulated by hTERT promoter and CMV enhancer can obviously and specifically inhibit and kill nasopharyngeal carcinoma cells in culture and nasopharyngeal carcinoma xenograft in nude mice in vivo, without obviously toxic side effects on nude mice. The targeted and enhanced TK gene vector with high performance may be a new tumour targeted gene therapy strategy clinically to aim directly at most malignant tumours including nasopharyngeal carcinoma, with more extensive anti-cancer spectrum.</p>

Enhancement of HepG2 cell radiosensitivity by mutant IkappaBalpha gene transfection / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the efficacy of transfection with a mutant IkappaBalpha gene (mIkappaBalpha) in enhancing the radiosensitivity of hepatocellular carcinoma cells.</p><p><b>METHODS</b>The hepatocellular carcinoma Hep G2 cells were divided into 3 group and transfected with the adenovirus containing mIkappaBalpha vector (Ad-mIkappaBalpha group), Ad vector (Adv group), or without treatment (parental control cells). Before and after irradiation of the cells with 6 Gy high-energy X ray, Western blotting was performed to measure the expression level of IkappaBalpha in the cytoplasm, and electrophoresis mobility shift assay (EMSA) carried out to evaluate the nuclear factor-kappaB (NF- kappa;B) activity in the cell nuclei, with the cell apoptosis detected using TUNEL assay. The radiosensitivity of the HepG2 cells were determined by comparison between the 3 groups in term of the surviving cell fractions (SF2) after 2-Gy X-ray exposure, and of the Do and Dq values obtained using the single-hit multi target model.</p><p><b>RESULTS</b>Before the X-ray exposure, the Hep G2 cells in Adv group and the control group showed low levels of IkappaBalpha absorbance in the cytoplasm, which were further decreased after the exposure. The NF-kappaB activity in the nuclei of the cells in the two groups was positive (+) before irradiation, and substantially enhanced (++) after the exposure and maintained the stably activated state. The apoptotic index of the cells in the two groups was 1.4 and 1.6 before irradiation, and increased to 8.9 and 11.7 after the irradiation, respectively. The cells in the Ad-mIkappaBalpha group, however, exhibited high levels of IkappaBalpha absorbance either before or after the irradiation, which were approximately 3 times that of the Adv group, but the NF-kappaB activity remained negative irrespective of the irradiation. The apoptotic index of the cells in the Ad-mIkappaBalpha group was 18.2 before irradiation, was increased to 88.3 after the irradiation. Among the 3 groups, Ad-mIkappaBalpha group had the smallest SF(2) value of 0.301 but the highest sensitivity enhancement ratio (SER) of2.99, with the lowest Do and Dq values (1.468 and 0.709, respectively).</p><p><b>CONCLUSION</b>mIkappaBalpha gene transfection into HepG2 cells inhibits the anti-apoptotic activity of NF-kappaB to enhance the radiosensitivity of the cells.</p>

Mutation detection of parkin gene by denaturing high performance liquid chromatography / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To detect parkin gene mutation of early-onset parkinsonism (EOP) by denaturing high performance liquid chromatography (DHPLC).</p><p><b>METHODS</b>The blood cell genomic DNA of 82 EOP patients was isolated. Exons of parkin gene were amplified by PCR. The PCR products were detected by DHPLC. The sample with abnormal peak shape was sequenced.</p><p><b>RESULTS</b>Three point mutations were identified in 82 EOP patients compared with 100 healthy controls. Mutations in intron include IVS1-39 G --> T and IVS9 +18 C --> T. The T1422C mutation was in coding region and resulted in 441 Cys --> Arg.</p><p><b>CONCLUSION</b>Three heterozygous mutations are found in sporadic EOP patients and genetic diagnosis of parkin gene by DHPLC is applicable in EOP patients.</p>

A novel method for detecting single-nucleotide polymorphisms by using ShineRoar probes / 中华检验医学杂志

Objective The present study demonstrates a novel,simple and cost-effective method for detecting known SNP genotyping by using ShineRoar probes.Methods The SNP of target genes detected by using the ShineRoar probes and melting curve analysis.Tumor necrosis factor receptor Ⅱ (TNFR Ⅱ) and apolipoprotein M (apoM) had been employed as target genes to describe the method in details.The PCR products of TNFR Ⅱ and apoM were collected and sequenced.Results The melting temperatures (TM) were significantly different between mutated genotypes and wild-type genotype.A biallelic SNP marker (T/ G) at position 196 in exon 6 of TNFR Ⅱ gene showed two melting valleys with the appropriate TMs at (52.84?0.75)℃ and (58.38?0.61)℃,respectively.For apoM T-778C,TMs of homozygous T genotype and C genotype were (42.55?0.73)℃ and (49.19?0.57)℃,respectively.Moreover,this genotyping method was validated by the DNA sequence analyses (Kappa=1,P=0.000).Conclusion It is concluded that this novel method is simple and economical and it is suitable for a large-scale genotyping screening.

An experimental animal model of chronic myocardial hibernation: comparative study of cine-MR, myocardial single photon emission computed tomography and pathology / 中华放射学杂志

Objective To establish the chronic low flow myocardial hibernation animal model in pigs, and to assess the diagnostic value for myocardial hibernation by using various imaging methods. Methods A total of 13 miniswine (30-40 kg) were used. All animals underwent general anesthesia and orotracheal intubation while the animals were mechanically ventilated. Under sterile conditions, left ventriculography and coronary angiography were performed by introduction of catheter into the right femoral artery. Further, a left anterolateral thoracotomy was performed in the third intercostal space. The proximal LCX was dissected free to allow placement of an ameroid constrictor. More than 1 month later, left ventriculography and coronary angiography were performed again, followed by cine MRI at rest and during stress with low dose of dobutamine (5 ?g?kg -1 ?min -1 ), respectively. Traditional and/or breath hold cine MRI were used to evaluate regional left ventricular wall motion, corresponding to basal, midventricular and apical short axis tomograms. Regional wall motion score index (WMSI) was calculated. At the same time 99m Tc MIBI myocardial SPECT was performed at rest and during nitroglycerin administration, respectively. All animals were finally sacrificed for pathological examination. Triphenyl tetrazolium chloride (TTC) staining was used to assess the myocardial infarction. Electron microscopy was used to identify myocardial cellular changes characteristic of hibernating myocardium. Results Three pigs died during surgery or within two weeks after surgery. One pig died of anesthesia during SPECT examination, 1 pig suffered from aneurysm, and another one pig showed negative findings. The other 7 pigs were found with hypokinetic ( n =4) or akinetic ( n =3) myocardial regions related to stenosed LCX (70%-99%). Resting cine MRI demonstrated decreased regional motion of the lateral and posteroinferior walls (ischemic regions) of the left ventricle ( n =7), compared with the nonischemic anteroseptal regions; but the low dose dobutamine (5 ?g?kg -1 ?min -1 ) could recover those hypokinetic or akinetic myocardial regions, characteristic of hibernating myocardium. Resting 99m Tc MIBI myocardial SPECT ( n =6) showed a fixed perfusion defect on the corresponding ischemic areas, which became reversible on the nitrate augmented myocardial perfusion imaging. It also indicated myocardial viability presented at the ischemic areas. TTC staining revealed patchy infarction of the area at risk localized to the endocardial surface ( n =3), and no myocardial infarction ( n =4). Electron microscopy of sections from the hibernating regions revealed loss of contractile materials, increased numbers of small mitochondria, and glycogen accumulation within viable cardiomyocytes, which had been described as hallmarks of hibernating myocardium. Conclusion Chronic low flow myocardial hibernation can be reproduced in an animal model during progressive coronary stenosis caused by ameroid constrictor.