VEGF-D-induced draining lymphatic enlargement and tumor lymphangiogenesis promote lymph node metastasis in a xenograft model of ovarian carcinoma.

BACKGROUND: Vascular endothelial growth factor (VEGF)-D has been shown to promote lymph node metastasis in several cancers. Although generally overexpressed in ovarian carcinoma, its role in nodal dissemination of this cancer is unclear. To clarify the role of VEGF-D and the underlying molecular mechanisms, we investigated the function of VEGF-D using a mouse xenograft model of ovarian cancer. METHODS: Human ovarian serous adenocarcinoma SKOV3 cells were transfected with VEGF-D recombinant plasmid DNA, or with control vectors. The cells were injected subcutaneously into the footpads of nude mice. Tumor growth was evaluated weekly. Draining lymphatics were observed grossly with Evan's blue lymphangiography. Tumoral lymphatics were delineated with both Evan's blue and LYVE-1 immunostaining. Tumor metastases to lymph nodes were evaluated by H&E and CA125/CD40 staining. Expression of VEGF-D in primary tumors and levels of CA125 in involved lymph nodes were examined by immunohistochemistry. Tumor cell apoptosis was analyzed by Hoechst dyeing. RESULTS: Mice bearing VEGF-D overexpressing xenografts showed a significantly higher rate of lymph node metastasis and markedly greater tumor volume compared with the controls. The functional lymphatic vessels were denser and enlarged in marginal and central tumor portions. Additionally, higher CA125 expression was observed in the involved lymph nodes. Mice bearing VEGF-D overexpressing xenografts also exhibited a markedly lower apoptotic index compared with the controls. CONCLUSIONS: Our data demonstrate the important role of VEGF-D in promoting lymph node metastasis by increasing tumor lymphangiogenesis, stimulating draining lymphatic vessel formation, and enhancing tumor invasiveness. Our findings show that VEGF-D can be a promising therapeutic target for ovarian cancer.

Individual patient data subgroup meta-analysis of surgery for spontaneous supratentorial intracerebral hemorrhage.

BACKGROUND AND PURPOSE: By 2010 there had been 14 published trials of surgery for intracerebral hemorrhage reported in systematic reviews or to the authors, but the role and timing of operative intervention remain controversial and the practice continues to be haphazard. This study attempted to obtain individual patient data from each of the 13 studies published since 1985 to better define groups of patients that might benefit from surgery. METHODS: Authors of identified published articles were approached by mail, e-mail, and at conferences and invited to take part in the study. Data were obtained from 8 studies (2186 cases). Individual patient data included patient's age, Glasgow Coma Score at presentation, volume and site of hematoma, presence of intraventricular hemorrhage, method of evacuation, time to randomization, and outcome. RESULTS: Meta-analysis indicated that there was improved outcome with surgery if randomization was [corrected] undertaken within 8 hours of ictus (P=0.003), or the volume of the hematoma was 20 to 50 mL (P=0.004), or the Glasgow Coma Score was between 9 and 12 (P=0.0009), or the patient was aged between 50 and 69 years (P=0.01). In addition, there was some evidence that more superficial hematomas with no intraventricular hemorrhage might also benefit (P=0.09). CONCLUSIONS: There is evidence that surgery is of benefit if undertaken early before the patient deteriorates. This work identifies areas for further research. Ongoing studies in subgroups of patients such as the Surgical Trial in Lobar Intracerebral Hemorrhage (STICH II) will confirm whether these interpretations can be replicated.

A novel transdermal plasmid-dimethylsulfoxide delivery technique for treatment of psoriasis.

BACKGROUND: Psoriasis is a chronic and relapsing inflammatory skin disease associated with various immunologic abnormalities. Repeated subcutaneous injection of interleukin-4 (IL-4) has been established as an effective treatment to counteract psoriasis. OBJECTIVE: We investigated whether gene therapy using IL-4 expression plasmid (pIL-4) via transdermal delivery was an alternative treatment for psoriasis. In our experiment, dimethylsulfoxide (DMSO) was used as a penetration enhancer. METHODS: At first, the penetration efficiency of the complex of reporter plasmid accompanied by DMSO was investigated both in vitro and in vivo. Then, the antipsoriasis efficiency of the treatment with pIL-4-DMSO was tested in mice. RESULTS: The expression of the reporter gene was detected in epidermis and dermis both in vitro and in vivo. More importantly, the psoriasis symptoms were relieved, and significant reductions in some psoriasis-associated factors were observed after pIL-4-DMSO treatment. CONCLUSION: We conclude that the topical application of pIL-4-DMSO can treat psoriasis to a significant extent.

Massive pulmonary haemorrhage due to severe trauma treated with repeated alveolar lavage combined with extracorporeal membrane oxygenation: A case report.

BACKGROUND: Massive pulmonary haemorrhage can spoil the entire lung and block the airway in a short period of time due to severe bleeding, which quickly leads to death. Alveolar lavage is an effective method for haemostasis and airway maintenance. However, patients often cannot tolerate alveolar lavage due to severe hypoxia. We used extracorporeal membrane oxygenation (ECMO) to overcome this limitation in a patient with massive pulmonary haemorrhage due to severe trauma and succeeded in saving the life by repeated alveolar lavage. CASE SUMMARY: A 22-year-old man sustained multiple injuries in a motor vehicle accident and was transferred to our emergency department. On admission, he had a slight cough and a small amount of bloody sputum; computed tomography revealed multiple fractures and mild pulmonary contusion. At 37 h after admission, he developed severe chest tightness, chest pain, dizziness and haemoptysis. His oxygen saturation was 68%. Emergency endotracheal intubation was performed, and a large amount of bloody sputum was suctioned. After transfer to the intensive care unit, he developed refractory hypoxemia and heparin-free venovenous ECMO was initiated. Fibreoptic bronchoscopy revealed diffuse and profuse blood in all bronchopulmonary segment. Bleeding was observed in the trachea and right bronchus, and repeated alveolar lavage was performed. On day 3, the patient's haemoptysis ceased, and ECMO support was terminated 10 d later. Tracheostomy was performed on day 15, and the patient was weaned from the ventilator on day 21. CONCLUSION: Alveolar lavage combined with ECMO can control bleeding in trauma-induced massive pulmonary haemorrhage, is safe and can be performed bedside.

Liposomal honokiol inhibits VEGF-D-induced lymphangiogenesis and metastasis in xenograft tumor model.

Lymph nodes metastasis of tumor could be a crucial early step in the metastatic process. Induction of tumor lymphangiogenesis by vascular endothelial growth factor-D may play an important role in promoting tumor metastasis to regional lymph nodes and these processes can be inhibited by inactivation of the VEGFR-3 signaling pathway. Honokiol has been reported to possess potent antiangiogenesis and antitumor properties in several cell lines and xenograft tumor models. However, its role in tumor-associated lymphangiogenesis and lymphatic metastasis remains unclear. Here, we established lymph node metastasis models by injecting overexpressing VEGF-D Lewis lung carcinoma cells into C57BL/6 mice to explore the effect of honokiol on tumor-associated lymphangiogenesis and related lymph node metastasis. The underlying mechanisms were systematically investigated in vitro and in vivo. In in vivo study, liposomal honokiol significantly inhibited the tumor-associated lymphangiogenesis and metastasis in Lewis lung carcinoma model. A remarkable delay of tumor growth and prolonged life span were also observed. In in vitro study, honokiol inhibited VEGF-D-induced survival, proliferation and tube-formation of both human umbilical vein endothelial cells (HUVECs) and lymphatic vascular endothelial cells (HLECs). Western blotting analysis showed that liposomal honokiol-inhibited Akt and MAPK phosphorylation in 2 endothelial cells, and downregulated expressions of VEGFR-2 of human vascular endothelial cells and VEGFR-3 of lymphatic endothelial cells. Thus, we identified for the first time that honokiol provided therapeutic benefit not only by direct effects on tumor cells and antiangiogenesis but also by inhibiting lymphangiogenesis and metastasis via the VEGFR-3 pathway. The present findings may be of importance to investigate the molecular mechanisms underlying the spread of cancer via the lymphatics and explore the therapeutical strategy of honokiol on antilymphangiogenesis and antimetastasis.

A promising cancer gene therapy agent based on the matrix protein of vesicular stomatitis virus.

The matrix (M) protein of vesicular stomatitis virus (VSV) plays a key role in inducing cell apoptosis during infection. To investigate whether M protein-mediated apoptosis could be used in cancer therapy, its cDNA was amplified and cloned into eukaryotic expression vector pcDNA3.1(+). The recombinant plasmid or the control empty plasmid pcDNA3.1(+) was mixed with cationic liposome and introduced into various tumor cell lines in vitro, including lung cancer cell LLC, A549, colon cancer cell CT26 and fibrosarcoma cell MethA. Our data showed that the M protein induced remarkable apoptosis of cancer cells in vitro compared with controls. Fifty micrograms of plasmid in a complex with 250 microg cationic liposome was injected intratumorally into mice bearing LLC or MethA tumor model every 3 days for 6 times. It was found that the tumors treated with M protein plasmid grew much more slowly, and the survival of the mice was significantly prolonged compared with the mice treated with the control plasmid. In MethA fibrosarcoma, the tumors treated with M protein plasmid were even completely regressed, and the mice acquired longtime protection against the same tumor cell in rechallenge experiments. Both apoptotic cells and CD8(+) T cells were widely distributed in M protein plasmid-treated tumor tissue. Activated cytotoxic T lymphocytes (CTLs) were further detected by means of (51)Cr release assay in the spleen of the treated mice. These results showed that M protein of VSV can act as both apoptosis inducer and immune response initiator, which may account for its extraordinary antitumor effect and warrant its further development in cancer gene therapy.

Humoral immunity responses against EGFR-positive tumor cells induced by xenogeneic EGFR expressed in the yeast Pichia pastoris.

The breaking of immune tolerance against self epidermal growth factor receptor (EGFR) should be a promising approach for the treatment of those receptor-positive tumors. We have previously shown that human EGFR as a xenoantigen induced a specific antitumor activity against EGFR-positive mouse tumors. Our further studies demonstrated that a recombinant form of extracellular domain of mouse EGFR provoked active cellular immunity responses against EGFR-positive human tumors. In this study, we investigated whether the recombinant murine EGFR expressed in the yeast Pichia pastoris would induce humoral immunity responses against EGFR-positive human tumors. To test this concept, polyclonal immunoglobulin (IgG), which was produced by vaccinating the rabbits with the recombinant mEGFR, was purified from the sera of the rabbits. We evaluated the antitumor activity of the polyclonal IgG in the nude mice bearing A431 tumors. Mice were i.v. treated with the purified IgG at 100 mg/kg 1 day before the mice were inoculated with the tumor cells and then twice per week for 4 weeks. Our results showed that the polyclonal IgG would efficiently inhibit the growth of the solid tumor in vivo. The antitumor effect of the polyclonal IgG may result from the increasing rate of apoptosis and induction of differentiation of the tumor cells in vivo. The present findings may provide insight into treatment of EGFR-positive tumors through induction of the humoral immunity responses based on xenogeneic homologous EGFR.

Human alpha-defensin-1 inhibits growth of human lung adenocarcinoma xenograft in nude mice.

Human alpha-defensin-1 (HNP1), a small antimicrobial peptide, shows cytotoxicity to tumor cells in vitro and inhibitory activity for pathologic neovascularization in vivo. Here, we did a gene therapy with a plasmid that expresses a secretable form of HNP1 for assaying its antitumor activity. The expression and secretion of HNP1 were determined by reverse transcription-PCR and ELISA in vitro. We found that expression of HNP1 in A549 tumor cells caused significant growth inhibition. This effect is most likely cell autonomous, as a significant amount of recombinant HNP1 protein was found to be accumulated in the cytoplasm by immunohistochemical staining using an anti-HNP1 antibody and the supernatant containing secreted HNP1 failed to produce any noticeable antitumor activity. Flow cytometry and Hoechst 33258 staining showed that the number of apoptotic cells among the A549 cells expressing recombinant HNP1 proteins was significantly greater than that of the nontransfected control cultures, suggesting that this growth-inhibitory activity was due to an apoptotic mechanism triggered by the intracellular HNP1. The antitumor activity of intracellularly expressed HNP1 was also shown in vivo. Decreased microvessel density and increased lymphocyte infiltration were observed in tumor tissue from HNP1-treated mice through histologic analysis. These results indicate that intracellularly expressed HNP1 induces tumor cell apoptosis, which inhibits tumor growth. The antiangiogenesis effect of HNP1 may contribute to its inhibitory activity in vivo, and HNP1 might involve the host immune response to tumor. These findings provide a rationale for developing HNP1-based gene therapy for cancer.

Immunity against tumor angiogenesis induced by a fusion vaccine with murine beta-defensin 2 and mFlk-1.

PURPOSE: Previous studies indicated that humoral or cellular immunity against murine vascular endothelial growth factor 2 (mFlk-1) was elicited to inhibit tumor growth. Here we describe a genetic fusion vaccine, pMBD2-mFlk-1, based on the targeting of a modified mFlk-1 to antigen-presenting cells by a murine beta-defensin 2 (MBD2) protein to induce both humoral and cellular immunity against mFlk-1, with the targeting especially focused on immature dendritic cells. EXPERIMENTAL DESIGN: The protective and therapeutic antitumor immunity of the fusion vaccine was investigated in mouse models. Antiangiogenesis effect was detected by immunohistochemical staining and alginate-encapsulate tumor cell assay. The mechanisms of the fusion vaccine were primarily explored by detection of autoantibodies and CTL activity and confirmed by the deletion of immune cell subsets. RESULTS: The fusion vaccine elicited a strong protective and therapeutic antitumor immunity through antiangiogenesis in mouse models, and this worked through stimulation of an antigen-specific CD8+ T-cell response as well as a specific B-cell response against mFlk-1. The findings were confirmed by depletion of immune cell subsets and in knockout mice. CONCLUSION: Our study showed that a fusion vaccine based on self immune peptide (MBD2) and self antigen (mFlk-1) induced autoimmunity against endothelial cells, resulting in inhibition of tumor growth, and could be further exploited in clinical applications of cancer immunotherapy.

Long-term stable expression of antisense cDNA of cyclin B1 profoundly inhibits the proliferation of tumor cells and suppresses tumorigenicity in implanted mice.

BACKGROUND: Cyclin B1 (CLB1) is necessary for mitotic initiation in mammalian cells and plays important roles in cancer development. Therefore, a potential strategy in cancer therapy is to suppress the activity of CLB1 by delivering antisense constructs of CLB1 into tumor cells. In previous CLB1 studies, antisense constructs with a short half life were often used and these constructs might not persistently inhibit CLB1. METHODS: We successfully created a recombinant plasmid encoding the full-length antisense cDNA of mouse cyclin B1 (AS-mCLB1) and transfected this construct to the murine Lewis lung carcinoma (LL/2) and CT-26 colon carcinoma (CT-26) cells. We isolated clones of LL/2 and CT-26 transfectants with stable expression of AS-mCLB1. Reverse transcriptional polymerase chain reaction (RT-PCR) and Western blot were applied to detect the expression of the mRNA and protein levels of CLB1. To further test the efficacy of this strategy in vivo, AS-mCLB1-expressing LL/2 and CT-26 transfectants were implanted into mice. RESULTS: We found the expression of the mRNA and protein levels of CLB1 decrease in these transfectants. The inhibition of CLB1 caused prominent G1 arrest, abnormal morphology, retarded cell growth and an increase in apoptosis. In AS-mCLB1-expressing LL/2 and CT-26 transfectants implanted mice, tumorigenicity was effectively suppressed compared with the controls. In addition, the expression of AS-mCLB1 also significantly increases the survival duration of implanted animals. CONCLUSION: AS-mCLB1 is likely to be useful in future cancer therapy, which may be associated with its ability to down-regulate the expression of CLB1 and then induce G1arrest and apoptosis in tumor cells.

[Researches on cloning and expression of the gene encoding Schistosoma japonicum tetraspanins extracellular loop 2 and its immunogenicity].

OBJECTIVE: To clone and express the gene (Sj-tsp-2) encoding extracellular loop 2 (EC-2) of tetraspanins (TSPs) of Schistosoma japonicum (Chinese strain), and then to study the antigenicity and immunogenicity of this protein. METHODS: Synthesized Sj-tsp-2 gene was cloned into prokaryotic expression vector pET32a to generate pET32a-Sj-tsp-2 recombinant plasmid, and transformed into competent E. coli BL21 (DE3). After inducing with IPTG, the expressed fusion protein was purified under nondenaturing conditions. RESULTS: The recombinant was confirmed by sequence analysis. The size of fusion protein and interest peptide was accords with the theoretical value by SDS-PAGE analysis. Additionally, the results of Western Blotting and ELISA demonstrated that the expressed protein had good immunogenicity. More importantly, we confirmed that TSP-2 is mainly located on the surface of S. japonicum. CONCLUSION: The successful expression and purification of recombinant protein Trx-Sj-TSP-2 will be very helpful for the further study of its protection role in animals.

Association between HLA-DR Expression and Multidrug-resistant Infection in Patients with Severe Acute Pancreatitis.

Multidrug-resistant (MDR) bacterial infection is a common complication of severe acute pancreatitis (SAP). This study aimed to explore the association between human leukocyte antigen-antigen D-related (HLA-DR) expression and multidrug-resistant infection in patients with SAP. A total of 24 SAP patients who were admitted to Nanjing Drum Tower Hospital between May 2015 and December 2016 were enrolled in the study. The percentages of CD4+, CD8+, natural killer (NK), and HLA-DR (CD14+) cells and the CD4+/CD8+ cell ratio on days 1,7,14, and 28 after admission were determined by flow cytometry. Eighteen patients presented with the symptoms of infection. Among them, 55.6% patients (10/18) developed MDR infection. The most common causative MDR organisms were Enterobacter cloacae and Acinetobacter baumannii. The CD4+/CD8+ cell ratio and the percentage of NK cells were similar between patients with non-MDR and patients with MDR infections. In patients without infection, the HLA-DR percentage was maintained at a high level throughout the 28 days. Compared to the patients without any infection, the HLA-DR percentage in patients with non-MDR infection was reduced on day 1 but increased and reached similar levels on day 28. In patients with MDR infection, the HLA-DR percentage remained below normal levels at all-time points. It was concluded that persistent down-regulation of HLA-DR expression is associated with MDR bacterial infection in patients with SAP.

Intraoperative real-time contrast-enhanced ultrasound angiography: a new adjunct in the surgical treatment of arteriovenous malformations.

OBJECT: The goal of this study was to combine the use of ultrasound contrast agents with intraoperative ultrasound techniques to identify intraoperatively a patient's vascular anatomy, including feeding arteries and draining veins of an intracranial arteriovenous malformation (AVM). METHODS: The authors examined 12 consecutive patients with AVMs that had been diagnosed on the basis of preoperative findings on magnetic resonance images and digital subtraction angiograms obtained between September 2003 and December 2005. After each patient had undergone a routine craniotomy, a bolus of contrast agent was injected intravenously, and a real-time microbubble perfusion process was observed to identify the feeding arteries and draining veins of the AVM in a single cross-section. The so-called burst-refill technique was used to sweep the lesion in multiple sections and orientations to obtain information on the surrounding vascular anatomy, after which the findings were compared with those obtained during preoperative imaging. RESULTS: Intraoperative ultrasonography provided high-quality images in every case. Although plain imaging failed to show an identifiable AVM boundary, color Doppler flow imaging clearly delineated the shape and margin of the AVM. Nevertheless, neither mode of imaging enabled the surgeons to categorically distinguish between feeding and draining vessels. The real-time perfusion process of microbubbles was first visualized 20 to 30 seconds after the SonoVue bolus injection, and the burst-refill technique made possible identification of the vascular anatomy of malformation lesions in multiple planes. CONCLUSIONS; Using both an ultrasound contrast agent and the burst-refill technique provided a rapid, convenient, and precise way of locating AVM feeding arteries intraoperatively. The combined technique seems warranted in the intraoperative treatment of AVMs.

A phase II clinical trial with cytotropic heterogeneous molecular lipids (CHML) for patients with hepatic malignancies.

Hepatocellular carcinoma (HCC) and other forms of metastatic liver cancer (MLC) have poor outcomes due to the limited treatment options. Surgery, radiotherapy and chemotherapy have a limited success. Thus, there is an urgent need for novel therapies for patients with advanced HCC and MLC. The response and toxicity profile of a novel biological anticancer agent, cytotropic heterogeneous molecular lipids (CHML), in 135 Asian patients with hepatic malignancies treated at five different hospitals in China from April 1998 to August 2003 is described. This trial included 97 patients with HCC and 38 with MLC. The majority of these patients had received conventional therapies and many had failed to respond or relapsed. CHML was administered by intra-arterial (i.a.) infusion with or without simultaneous intravenous (i.v.) infusion for 25 days with a rest of 2-4 weeks between each cycle. Fifty three percent of patients received two cycles, and 47% received three cycles. The complete response (CR) rates were 23% for HCC and 29% for MLC with an overall CR of 24%. The overall partial response (PR) was 53%. The patients with earlier stages and limited tumor burden had a better response, but a few patients with advanced disease also achieved PR. The patients who achieved CR or PR had a significant increase in long-term survival for up to five years. The treatment with CHML resulted in minimal toxicity and the reported adverse reactions were not higher than grade II. CHML is an effective therapy for hepatic malignancies, showing responses and increases in survival in patients in whom other therapies have failed. CHML is well tolerated and is an excellent candidate for Phase III clinical trials.

[Clinical experience of 70 cases of cerebral arteriovenous malformations embolization with Onyx, a novel liquid embolic agent].

OBJECTIVE: To report our clinical experience of using Onyx, a new liquid embolic agent, to treat cerebral arteriovenous malformations (AVMs) as well as its efficacy. METHODS: Seventy cases were placed with 6F sheath in the femoral artery after Seldinger puncture and 6F guiding catheter was introduced into the internal carotid artery or vertebral artery, then a microcatheter was navigated into the nidus of AVMs. Slow injection of Onyx under fluoroscopic control was performed to embolize cerebral AVMs using the "plug and push" technique. RESULTS: Thirteen AVM cases (18.6%) were totally occluded by Onyx and 5 cases of which didn't recurrence at 6-month after operation. Thirty-eight cases (54.3%) were subtotally occluded, while another 19 cases (27.1%) were partially embolized. Severe cerebral hemorrhage occurred in 4 cases, 2 of which had mild to severe hemiplegia after operation, and one died. Mild hemiplegia was also found in 1 case due to functional area embolization, and visual field deficit in 2 cases. CONCLUSIONS: Onyx has unique and distinctive superiority in treating cerebral AVMs. Nonetheless, the correct embolization technique should be learned to achieve good clinical results and avoid complications.

Effects of ischemic preconditioning on blood-brain barrier permeability and MMP-9 expression of ischemic brain.

The study was designed to investigate the effects of ischemic preconditioning (IP) on permeability of blood-brain barrier (BBB) and expression of matrix metalloproteinase-9 (MMP-9) in subsequent ischemic hemisphere. Rats were divided into four groups, one group was used as control, and the other three groups were given three different pretreatments: the first group received a saline injection into the right internal carotid artery (SI), the second group underwent both left and right carotid arteries occlusion (BCAO), and the third group was treated with BCAO and SI simultaneously (BS). After 24 hours of pretreatments, the focal cerebral ischemia was induced by inserting a thread into the right middle cerebral artery causing occlusion (MCAO). Brain water content, BBB permeability and MMP-9 expression of ischemic hemisphere brains were measured at 24 and 48 hours after MCAO. After 24 and 48 hours MCAO, averages for brain water content were 82.92 and 83.12% in BS group, 85.19 and 85.73% in SI group and 86.06 and 85.88% in BCAO group. Evans blue content of ischemic hemispheres were 14.01 and 11.74 microg/mm(3) at 24 and 48 hours after MCAO in BS group, which were lower than the other two groups, 16.22, 15.01 and 16.61, 15.58 microg/mm(3), respectively (p<0.01). The expression levels of MMP-9 in ischemic hemisphere in BS were lower than that in other two groups (p<0.01). Therefore, ischemic preconditioning could ameliorate brain edema and BBB disruption caused by subsequent cerebral ischemia. Ischemic preconditioning could decrease MMP-9 protein and mRNA expression, which may be an important mechanism of cerebral ischemic tolerance.

A study on blood-brain barrier ultrastructural changes induced by cerebral hypoperfusion of different stages.

OBJECTIVES: To evaluate the ultrastructural changes in the blood-brain barrier (BBB) induced by cerebral hypoperfusion of different stages, which may predispose the brain to the formation of vasogenic edema and hemorrhage under cerebral luxury perfusion. METHODS: Twenty cerebral steal models with left neck arteriovenous fistula (AVF) were surgically created in Wistar's rats, leading to a noninfarctional reduction in the cerebral blood flow (CBF) by between 25 and 50%, resulting in cerebral hypoperfusion in the AVF side for 3 days (acute stage), 3 weeks (subacute stage) and 3 months (chronic stage), respectively. Another six sham-operated models were made in age-matched rats as control. The BBB ultrastructural changes were assessed by transmission electron microscopy. Ridit analysis was conducted to compare the positive ratio of ultrastructural changes among multiple groups. RESULTS: Electron microscopy demonstrated no ultrastructural change at the acute stage, however, at the subacute stage, slight vacuolar degeneration was found in the astrocytic foot process layer encircling the capillaries; furthermore, at the chronic stage, the astrocytic foot processes expressed marked vacuolization associated with the adjacent astrocytic degeneration. Meanwhile, in both capillary endothelium and basal lamina layers, no abnormal ultrastructures similar to those in the astrocytic foot processes layer were identified. After cerebral luxury perfusion took place, BBB was disrupted where astrocytic foot processes vacuolization was most distinguished. CONCLUSION: Astrocytes generate ultrastructural abnormality as a result of chronic cerebral hypoperfusion. Astrocytic foot process vacuolization, which constitutes the major ultrastructural change in the BBB, is the extension of the degeneration of astrocyte body. It is inferred that BBB is prone to structure weakness and function instability, which forms the morphological basis of cerebral luxury perfusion.

Risk factors for intracranial aneurysm in a Chinese ethnic population.

BACKGROUND: Intracranial aneurysm (IAN) is a protruding bubble or a sac on a brain artery that balloons out over time, which may lead to spontaneous subarachnoid hemorrhage (SAH), ultimately disability and mortality. Current research indicates that the disease is due to multiple causes, including environmental factors and various congenital abnormalities of blood vessels. Apart from congenital predisposition, various high-risk factors such as sex, age, hypertension, and atherosclerosis are involved in the formation of intracranial aneurysms. The aim of this study was to investigate the risk factors associated with the formation of sporadic intracranial aneurysms in Chinese Han ethnic patients. METHODS: A total of 251 patients with intracranial aneurysm and 338 patients with other cerebral diseases (control group) were enrolled in this study. Single factor and logistic regression model were used to analyze the association of intracranial aneurysms with age; sex; cigarette smoking; alcohol or cocaine consumption; history of hypertension, coronary artery disease, diabetes mellitus and inherited connective tissue disease; and the levels of fasting blood glucose and blood fat. The data expressed as mean +/- standard deviation were processed with the statistical software SPSS13. Quantitative and qualitative data were analyzed by the independent-sample t test, and the chi-square test respectively. Logistic regression method was used to analyze the multiple factors. RESULTS: In the 251 patients, 163 (64.94%) were at age of 40 to 60 years. Sex (OR, 1.41; 95% CI, 1.01 - 1.96), cigarette smoking (OR, 1.81; 95% CI, 1.06 - 3.10), hypertension (OR, 2.32; 95% CI, 1.30 - 4.16) and fasting blood glucose were significantly associated with intracranial aneurysm (P < 0.05). Intracranial aneurysm was correlated with alcohol consumption, coronary artery disease, and the level of blood lipids (P > 0.05). Using logistic regression analysis, we identified female sex and advanced age as significant risk factors for sporadic intracranial aneurysms. CONCLUSIONS: Sporadic intracranial aneurysms mostly occur in people aged 40 to 60 years. Feminine, cigarette smoking, and hypertension are independent risk factors for the disease, and the gender is the most significant factor. Advanced age can increase the effect of these risk factors.

[Construction of pORF-mIG and activity evaluation of MIG in vitro].

OBJECTIVE: To establish a transformant of monokine induced by interferon-gamma (MIG) with the eukaryotic expression vector for further investigating the efficacy of its use in antitumor gene therapy. METHODS: The MIG full-length cDNA was amplified from pBLAST-MIG and was cloned into the eukaryote expression vector pORF-mcs, and the resulted recombinant plasmid was named pORF-MIG. The recombinant pORF-MIG was determined with restriction enzyme and sequencing, and then it was transfected into COS-7 cells by Lipfectamin. Expression of the transformant was detected by immunoblot assay, and the bioactivity was assessed by chemotaxis assay. RESULTS: The restriction analysis and the sequence determination confirmed that the recombinant pORF-MIG contained the MIG full-length cDNA. And the transformants of pORF-MIG expressed the MIG protein which could apparently attract the activated lymphocytes. CONCLUSION: The recombinant pORF-MIG was constructed successfully, and this recombinant eukaryotic expression vector could be used in additional studies on the biological effect of MIG and its use in anti-tumor gene therapy.

Antitumor activity of pluripotent cell-engineered vaccines and their potential to treat lung cancer in relation to different levels of irradiation.

Cancer stem cells (CSCs) are critical for tumor initiation/maintenance and recurrence or metastasis, so they may serve as a potential therapeutic target. However, CSC-established multitherapy resistance and immune tolerance render tumors resistant to current tumor-targeted strategies. To address this, renewable multiepitope-integrated spheroids based on placenta-derived mesenchymal stem cells (pMSCs) were X-ray-modified, at four different irradiation levels, including 80, 160, 240, and 320 Gy, as pluripotent biologics, to inoculate hosts bearing Lewis lung carcinoma (LL2) and compared with X-ray-modified common LL2 cells as control. We show that the vaccines at the 160/240 Gy irradiation levels could rapidly trigger tumor cells into the apoptosis loop and evidently prolong the tumor-bearing host's survival cycle, in contrast to vaccines irradiated at other levels (P<0.05), with tumor-sustaining stromal cell-derived factor-1/CXCR4 pathway being selectively blockaded. Meanwhile, almost no or minimal toxicity was detected in the vaccinated hosts. Importantly, 160/240 Gy-irradiated vaccines could provoke significantly higher killing of CSCs and non-CSCs, which may provide an access to developing a novel biotherapy against lung carcinoma.