Construction of specific Smo lentivirus and expression of infected pancreatic cancer cells positive for CD24CD44 surface antibody.

This study aimed to construct a vector lentivirus carrying the Smo gene and transfect pancreatic cancer cells positive for CD24CD44 surface antibody and detect the infectivity. A lentivirus carrying a specific Smo fragment was designed and synthesized, and its functionality was tested. An overexpression group, inhibitory group, and negative control group were used for subsequent experimental research and comparison. A virus was successfully designed and produced. The best viral load was the 1X106 TU virus, where the cell growth and fluorescence effect of culture wells with polybrene dilution were the best. These are the transfection conditions and transfection param-eters for subsequent experiments. This plasmid was detected with a flag antibody by Western blot. The result was that it had a large specific 250kD band, and the membrane protein was overexpressed successfully. The expression results of Smo in five groups of cells after virus transfection detected by RT-PCR: blank group were 1.0038±0.0344, CON238 negative group: 1.0276±0.2944d, CON077 negative group: 0.8793±0.0402; LV-SMO15570-2 overexpres-sion group: 2.7479±0.8308, and LV-SMO-RNAi37304-1 inhibition group: 0.2386±0.0481. There were differences among the overexpression group and inhibition group with the other three groups. Homogeneity of variance: Bartlett F = 4.3530, P = 0.0016 < 0.05, heterogeneous. K-W test: cc2 = 10.9905* P = 0.0267, and there was a statisti-cally significant difference. The designed virus achieved the goal requirements. An sRNA fragment was designed for the key gene Smo of the Hh signaling pathway, and a vector lentivirus carrying this fragment was successfully constructed. The expression of Smo was analyzed after transfecting SW1990CD24CD44 positive cells, suggesting that the function of the RNA fragment designed for the key gene Smo in this experiment was successful.

CD28 deficiency attenuates primary blast-induced renal injury in mice via the PI3K/Akt signalling pathway.

INTRODUCTION: Primary blast affects the kidneys due to direct shock wave damage and the production of proinflammatory cytokines without effective treatment. CD28 has been reported to be involved in regulating T cell activation and secretion of inflammatory cytokines. The aim of this study was to investigate the influence of primary blast on the kidney and the effect of CD28 in mice. METHODS: A mouse model of primary blast-induced kidney injury was established using a custom-made explosive device. The severity of kidney injury was investigated by H&E staining. ELISA was applied to study serum inflammation factors' expression. Western blot assays were used to analyse the primary blast-induced inflammatory factors' expression in the kidney. Immunofluorescence analysis was used to examine the PI3K/Akt signalling pathway. RESULTS: Histological examination demonstrated that compared with the primary blast group, CD28 deficiency caused a significant decrease in the severity of the primary blast-induced renal injury. Moreover, ELISA and western blotting revealed that CD28 deficiency significantly reduced the levels of interleukin (IL)-1ß, IL-4 and IL-6, and increased the IL-10 level (p<0.05). Finally, immunofluorescence analysis indicated that PI3K/Akt expression also changed. CONCLUSIONS: CD28 deficiency had protective effects on primary blast-induced kidney injury via the PI3K/Akt signalling pathway. These findings improve the knowledge on primary blast injury and provide theoretical basis for primary blast injury treatment.

Specific Heat Study of 1D and 2D Excitations in the Layered Frustrated Quantum Antiferromagnets Cs_{2}CuCl_{4-x}Br_{x}.

We report an experimental and theoretical study of the low-temperature specific heat C and magnetic susceptibility χ of the layered anisotropic triangular-lattice spin-1/2 Heisenberg antiferromagnets Cs_{2}CuCl_{4-x}Br_{x} with x=0, 1, 2, and 4. We find that the ratio J^{'}/J of the exchange couplings ranges from 0.32 to ≈0.78, implying a change (crossover or quantum phase transition) in the materials' magnetic properties from one-dimensional (1D) behavior for J^{'}/J<0.6 to two-dimensional (2D) behavior for J^{'}/J≈0.78. For J^{'}/J<0.6, realized for x=0, 1, and 4, we find a magnetic contribution to the low-temperature specific heat, C_{m}∝T, consistent with spinon excitations in 1D spin-1/2 Heisenberg antiferromagnets. Remarkably, for x=2, where J^{'}/J≈0.78 implies a 2D magnetic character, we also observe C_{m}∝T. This finding, which contrasts the prediction of C_{m}∝T^{2} made by standard spin-wave theories, shows that Fermi-like statistics also plays a significant role for the magnetic excitations in spin-1/2 frustrated 2D antiferromagnets.

Quantum Criticality of an Ising-like Spin-1/2 Antiferromagnetic Chain in a Transverse Magnetic Field.

We report on magnetization, sound-velocity, and magnetocaloric-effect measurements of the Ising-like spin-1/2 antiferromagnetic chain system BaCo_{2}V_{2}O_{8} as a function of temperature down to 1.3 K and an applied transverse magnetic field up to 60 T. While across the Néel temperature of T_{N}∼5 K anomalies in magnetization and sound velocity confirm the antiferromagnetic ordering transition, at the lowest temperature the field-dependent measurements reveal a sharp softening of sound velocity v(B) and a clear minimum of temperature T(B) at B_{⊥}^{c,3D}=21.4 T, indicating the suppression of the antiferromagnetic order. At higher fields, the T(B) curve shows a broad minimum at B_{⊥}^{c}=40 T, accompanied by a broad minimum in the sound velocity and a saturationlike magnetization. These features signal a quantum phase transition, which is further characterized by the divergent behavior of the Grüneisen parameter Γ_{B}∝(B-B_{⊥}^{c})^{-1}. By contrast, around the critical field, the Grüneisen parameter converges as temperature decreases, pointing to a quantum critical point of the one-dimensional transverse-field Ising model.

Inhibition of pulmonary carcinoma proliferation or metastasis of miR-218 via down-regulating CDCP1 expression.

OBJECTIVE: Pulmonary carcinoma is one common malignant tumor with a high risk of recurrence and metastasis. Non-small cell lung cancer (NSCLC) is the most common subtype. As one tumor biomarker, microRNA (miR) has tissue sensitivity and can facilitate oncogene or inhibit tumor suppressor gene. MiR-218 has abnormal expression and can work as one molecular marker for tumors. However, its expression and function mechanism in lung cancer cells have not been fully illustrated. MATERIALS AND METHODS: In vitro cultured pulmonary adenoma A549 cells and normal bronchial epithelial cell line 16HBE were tested for miR-218 expression. A549 cells were transfected with miR-218 mimic or negative controls, followed by real-time PCR quantifying for miR-218. MTT method was used to test cell proliferation, whilst Transwell chamber was adopted for measuring cell invasion. Dual luciferase reporter gene assay (DLRGA) was used to test target relationship between miR-218 and CDCP1. Western blot was used to test CDCP1 expression. RESULTS: MiR-218 was down-regulated in A549 cells compared to 16HBE (p<0.05). Transfection of miR-218 mimic significantly facilitated miR-218 expression, inhibited tumor proliferation or invasion. As the target gene of miR-218, CDCP1 expression was suppressed by miR-218 over-expression (p<0.05 compared to control group). CONCLUSIONS: MiR-218 inhibits NSCLC proliferation or metastasis via down-regulating CDCP1, and can work as one novel molecular target for lung cancer diagnosis.

Establishment, Differentiation, Electroporation and Nuclear Transfer of Porcine Mesenchymal Stem Cells.

The limited success of somatic cell nuclear transfer (SCNT) is largely attributed to defects in epigenetic reprogramming of the donor genome. Donor cell types with distinct potential competence may offer different epigenetic flexibility for subsequent genome reprogramming in SCNT. Stem cells possibly enable their genomes to be more readily reprogrammed than differentiated cells. To improve the efficiency of cloning, porcine mesenchymal stem cells (pMSCs) were isolated and well identified by 6-channel flow cytometry and differentiation assays and were used as donors in SCNT. Compared with porcine embryonic fibroblasts (pEFs), our results showed that pMSCs markedly enhanced cloned embryo development in terms of cleavage and blastocyst formation (p < 0.05). To enhance the epigenetic flexibility of pMSCs, classical reprogramming factors (RFs) were transfected by electroporation, and we achieved optimization with ectopic expression of RFs in pMSCs. Our results suggest that the epigenetic status of donor cells has an improvement on genome reprogramming, and multipotent pMSCs favoured subsequent embryonic development.

A chronic generalized bi-directional brain-machine interface.

A bi-directional neural interface (NI) system was designed and prototyped by incorporating a novel neural recording and processing subsystem into a commercial neural stimulator architecture. The NI system prototype leverages the system infrastructure from an existing neurostimulator to ensure reliable operation in a chronic implantation environment. In addition to providing predicate therapy capabilities, the device adds key elements to facilitate chronic research, such as four channels of electrocortigram/local field potential amplification and spectral analysis, a three-axis accelerometer, algorithm processing, event-based data logging, and wireless telemetry for data uploads and algorithm/configuration updates. The custom-integrated micropower sensor and interface circuits facilitate extended operation in a power-limited device. The prototype underwent significant verification testing to ensure reliability, and meets the requirements for a class CF instrument per IEC-60601 protocols. The ability of the device system to process and aid in classifying brain states was preclinically validated using an in vivo non-human primate model for brain control of a computer cursor (i.e. brain-machine interface or BMI). The primate BMI model was chosen for its ability to quantitatively measure signal decoding performance from brain activity that is similar in both amplitude and spectral content to other biomarkers used to detect disease states (e.g. Parkinson's disease). A key goal of this research prototype is to help broaden the clinical scope and acceptance of NI techniques, particularly real-time brain state detection. These techniques have the potential to be generalized beyond motor prosthesis, and are being explored for unmet needs in other neurological conditions such as movement disorders, stroke and epilepsy.

Effects of cholesterol on CCK-1 receptors and caveolin-3 proteins recycling in human gallbladder muscle.

The contraction of gallbladders (GBs) with cholesterol stones is impaired due to high cholesterol concentrations in caveolae compared with GBs with pigment stones. The reduced contraction is caused by a lower cholecystokinin (CCK)-8 binding to CCK-1 receptors (CCK-1R) due to caveolar sequestration of receptors. We aimed to examine the mechanism of cholesterol-induced sequestration of receptors. Muscle cells from human and guinea pig GBs were studied. Antibodies were used to examine CCK-1R, antigens of early and recycling endosomes, and total (CAV-3) and phosphorylated caveolar-3 protein (pCAV-3) by Western blots. Contraction was measured in muscle cells transfected with CAV3 mRNA or clathrin heavy-chain small-interfering RNA (siRNA). CCK-1R returned back to the bulk plasma membrane (PM) 30 min after CCK-8 recycled by endosomes, peaking at 5 min in early endosomes and at 20 min in recycling endosomes. Pretreatment with cholesterol-rich liposomes inhibited the transfer of CCK-1R and of CAV-3 in the endosomes by blocking CAV-3 phosphorylation. 4-Amino-5-(4-chloro-phenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (inhibitor of tyrosine kinase) reproduced these effects by blocking pCAV-3 formation, increasing CAV-3 and CCK-1R sequestration in the caveolae and impairing CCK-8-induced contraction. CAV-3 siRNA reduced CAV-3 protein expression, decreased CCK-8-induced contraction, and accumulated CCK-1R in the caveolae. Abnormal concentrations of caveolar cholesterol had no effect on met-enkephalin that stimulates a delta-opioid receptor that internalizes through clathrin. We found that impaired muscle contraction in GBs with cholesterol stones is due to high caveolar levels of cholesterol that inhibits pCAV-3 generation. Caveolar cholesterol increases the caveolar sequestration of CAV-3 and CCK-1R caused by their reduced recycling to the PM.

Inhaled ultrafine particulate matter affects CNS inflammatory processes and may act via MAP kinase signaling pathways.

In addition to evidence that inhalation of ambient particulate matter (PM) can increase cardiopulmonary morbidity and mortality, the brain may also constitute a site adversely effected by the environmental presence of airborne particulate matter. We have examined the association between exposure to PM and adverse CNS effects in apolipoprotein E knockout (ApoE-/-) mice exposed to two levels of concentrated ultrafine particulate matter in central Los Angeles. Mice were euthanized 24h after the last exposure and brain, liver, heart, lung and spleen tissues were collected and frozen for subsequent bioassays. There was clear evidence of aberrant immune activation in the brains of exposed animals as judged by a dose-related increase in nuclear translocation of two key transcription factors, NF-kappaB and AP-1. These factors are involved in the promotion of inflammation. Increased levels of glial fibrillary acidic protein (GFAP) were also found consequent to particulate inhalation suggesting that glial activation was taking place. In order to determine the mechanism by which these events occurred, levels of several MAP kinases involved in activation of these transcription factors were assayed by Western blotting. There were no significant changes in the proportion of active (phosphorylated) forms of ERK-1, IkB and p38. However, the fraction of JNK in the active form was significantly increased in animals receiving the lower concentration of concentrated ambient particles (CAPs). This suggests that the signaling pathway by which these transcription factors are activated involves the activation of JNK.

Effects of fusion/activation methods on development of embryos produced by nuclear transfer of porcine fetal fibroblast.

The present studies were carried out to investigate the effects of intensity of dc pulse, number of dc pulse and equilibration before fusion/activation on developmental ability of porcine embryos derived from nuclear transfer. In experiment 1, different fusion/activation intensity (two dc pulses of 0.4, 0.8, 1.2, 1.6 and 2.0 kV/cm for 30 micros, respectively) was carried out to investigate development of embryos. In experiment 2, the reconstructed oocytes were fused and activated with one, two or three dc pulses of 1.2 kV/cm for 30 micros. In experiment 3, reconstructed oocytes were equilibrated in TCM-199 medium for 0-6 h, respectively, and fused/activated with one dc pulse of 1.2 kV/cm for 30 micros. The reconstructed embryos were cultured in PZM-3 medium containing 0.3% BSA. When oocytes were fused with donor cell by two dc pulses of 0.4 kV/cm for 30 micros, the rates of cleavage and blastocyst formation were significantly lower (32.9% and 2.5%) than those of fused by 0.8 kV/cm (59.0% and 17.4%) or 1.2 kV/cm (63.3% and 18.4%), respectively. One dc pulse of 1.2 kV/cm for 30 micros was enough to fuse and activate embryos to develop to blastocyst (24.8%). Equilibration for 2-3 h in TCM-199 before fusion/activation was beneficial for improving the developmental ability of embryos produced by nuclear transfer (25.6-23.3% at blastocysts).

Chronic exposure to low levels of aluminum alters cerebral cell signaling in response to acute MPTP administration.

Two-month-old male B/6C3F1 mice were treated for 10 weeks with 100 microM aluminum lactate (Al) in drinking water. This dose of Al did not alter body weight, and there was no evidence of systemic toxicity. The degree of phosphorylation of several kinases which lead to transcription factor activation (reflecting the extent of their activation) was studied. The proportion of extracellular signal-regulated kinase (ERK) that was activated was depressed in cortex but not in the hippocampus following treatment but c-Jun N-terminal kinase (JNK), p38, IkappaB phosphorylation was unaltered in either tissue. Treatment of mice with 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) alone produced no significant changes in the degree of activation of any transcription factor studied. When MPTP dosing had been preceded by extended exposure to low levels of Al in drinking water, ERK activation was profoundly depressed in cortex and hippocampus, whereas JNK in hippocampus and IkappaB in cortex were greatly elevated. These changes consequent to exposure to both Al and MPTP were accompanied by an increase in NF-kappaB in both regions, whereas AP-1 was elevated in the hippocampus alone. Neither agent alone modulated AP-1 or NF-kappaB. Thus a synergistic interaction occurred between the toxicants. This interaction tended to promote the functioning of a kinase largely associated with inflammation and to depress that of ERK, which is associated with maintenance of cell survival. It is concluded that exposure to levels of Al with no evident toxicity can worsen the response to an acute challenge with MPTP. Al treatment alone was able to increase striatal 3,4-dihydroxyphenylacetic acid levels, suggesting an elevation of the rate of dopamine turnover in the striatum. However, no interaction in alteration of monoamine levels was found between Al and MPTP.

A steroidal saponin from the seeds of Allium tuberosum.

A steroidal saponin, named tuberoside, together with seven known compounds, were isolated from the seeds of Allium tuberosum Rottl. ex Spreng. Its structure was established by spectroscopic data, hydrolysis, and comparison with spectral data of known compounds to be (2alpha, 3beta, 5alpha, 25S)-2,3,27-trihydroxyspirostane 3-O-alpha-L-rhamnopyranoyl-(1-->2)-O-[alpha-L-rhamnopyranoyl-(1-->4)]-beta-D-glucopyranoside.

History of Chinese stereotactic and functional neurosurgery.

Chinese stereotactic and functional neurosurgery started in 1963. Dr. Jian-Ping Xu did stereotactic surgery for Parkinson's disease with a small Cartesian coordinate stereotactic device which he designed. In 1983, the first Chinese Institute of Stereotactic and Functional Neurosurgery was established by Dr. Jian-Ping Xu and Dr. Ye-Han Wang in the Anhui Provincial Hospital in the city of Hefei. Since then, the Institute has hosted an annual National Workshop on Stereotactic and Functional Neurosurgery, where more than 80% of the functional neurosurgeons now practicing in China have been trained. In 1986, the Chinese Society of Stereotactic and Functional Neurosurgery was established, and the first issue of the Chinese Journal of Stereotactic and Functional Neurosurgery was published. With more than 35 years of development, stereotactic and functional neurosurgery has become a very important branch of surgery in China. More than 5,000 functional neurosurgery procedures and more than 8,000 stereotactic radiosurgery procedures are now performed annually.

[Preoperative superselective intra-internal iliac arterial chemotherapy for invasive bladder cancer and curative analysis].

OBJECTIVE: To study clinical effects of preoperative intra-internal iliac-arterial chemotherapy for invasive bladder cancer. METHODS: 14 cases of invasive bladder cancer underwent superselective intra-internal iliac-arterial chemotherapy before conservative cystectomy. RESULTS: After 2-3 week therapy, reduction of cancer volume (20%-50%), clotted by thrombus in the blood vessels of the cancer or circumferential tissue, and decreased bleeding were found. Pathomorphological features included large patchy necrosis, degeneration, and inflammatory changes of carcinoma tissue. Follow-up ranged from 5 to 42 months (mean 19.6 months). One T3 N1 M0) of the cases (0.7%) relapsed into T1N0M0 and the others did not. CONCLUSIONS: The intra-internal iliac-arterial method, simple, safe, coordinates in pre-operation for invasive bladder cancer, and increases the operative opportunity for preservation of functional bladder.

Combinatorial discovery of oxidative dehydrogenation catalysts within the Mo-V-Nb-O system.

Combinatorial methodologies were used for the synthesis and screening of mixed metal oxide heterogeneous catalysts. Primary screening at low reactant conversions at a throughput of greater than 10,000 catalyst compositions per month was performed by using simultaneous MS and photothermal deflection spectroscopy on spatially separated thick film catalysts with approximately 200 microg per catalyst prepared by using automated liquid dispensing. Secondary screening under realistic operating conditions was performed at a throughput of greater than 3,000 catalyst compositions per month on approximately 50 mg of catalyst in an array of fixed bed microreactors with gas chromatograph detection. The approach was validated by the discovery of catalysts with superior performance to those previously described for the oxidative dehydrogenation of ethane to ethylene. We show the full implementation and integration of combinatorial methodologies for synthesis, screening, discovery, and optimization of multicomponent heterogeneous catalysts.

Short Communication: An apospory-specific genomic region is conserved between Buffelgrass (Cenchrus ciliaris L.) and Pennisetum squamulatum Fresen.

Twelve molecular markers linked to pseudogamous apospory, a form of gametophytic apomixis, were previously isolated from Pennisetum squamulatum Fresen. No recombination between these markers was found in a segregating population of 397 individuals (Ozias-Akins et al. 1998, Proc. Natl Acad. Sci. USA, 95, 5127-5132). The objective of the present study was to test if these markers were also linked to the aposporous mode of reproduction in two small segregating populations of Cenchrus ciliaris (= Pennisetum ciliare (L.)Link), another apomictic grass species. Among 12 markers (sequence characterized amplified regions, SCARs), six were scored as dominant markers between aposporous and sexual C. ciliaris genotypes (presence/absence, respectively). Five were always linked to apospory and one showed a low level of recombination in 84 progenies. Restriction fragment length polymorphisms (RFLPs) were observed between sexual and apomictic phenotypes for three of the six remaining SCARs from P. squamulatum when used as probes. No recombination was observed in the F1 progenies. Preliminary data from megabase DNA analysis and sequencing in both species indicate that an apospory-specific genomic region (ASGR) is highly conserved between the two species. Although C. ciliaris has a smaller genome size to P. squamulatum, a higher copy number for markers linked to apospory found in the former may impair the progress of positional cloning of gene(s) for apomixis in this species.

Sphingosine derivatives from the seeds of Allium tuberosum.

A new ceramide, named tuber-ceramide (2), along with a known cerebroside (1) were isolated from the seeds of Allium tuberosum. The structure of tuber-ceramide was determined on the basis of spectral data as N-(2',3'-dihydroxy-tetracosenoyl)-2-amino-1,3,4-trihydroxy octadecane (2). This is the first report of sphingosine derivatives isolated from the genus Allium.

[Chemical constituents of fructus Amomi].

Seven compounds have been isolated from Fructus Amomi. They were identified as bornyl acetate, camphor, borneol, beta-sitosterol, vanillic acid, stearic acid and palmitic acid. The vanillic acid was found in this medicine for the first time. Fifty-seven components were identified from the volatile oil of Fructus Amomi on the basis of GC-MS analysis. Eight components in the volatile oil over 1% in content are bornyl acetate, camphor, borneol, limonene, camphene, myrecene, carene-3 and alpha-terpeneol.

[Studies on the steroidal alkaloids from Sarcococca vagans].

The root of Sarcococca vagans Stapt (Buxaceae) is well-known Chinese folk drug called as "Jie Gu Mu", distributed in southern of China. They are used in treatment of traumatic injury. Four new steroidal alkaloids, named sarcovagine A, B, C and D, were isolated from the root of Sarcococca vagans. Their structures were established on the basis of spectral evidence.