Lac/Tet dual-inducible system functions in mammalian cell lines

The Escherichia coli Lac repressor (Lac system) and tetracycline responsive promoter (Tet system) systems have been used individually to regulate gene expression at the cellular as well as the organismal levels. In this study, these two systems were combined (designated Lac/Tet dual-inducible system) to regulate two inducible genes simultaneously in a single cell. The isopropyl-beta-D-thiogalactopyranoside (IPTG) and tetracycline (used for the operation of the Lac and the Tet systems) were non-cytotoxic to the cells when added together into the cells at around the optimal concentrations (IPTG: < or = 5 mM; tetracycline: < 1.5 micrograms). The rate and efficiency of induction and repression of two inducible genes regulated by the Lac/Tet dual-inducible system were similar to the results obtained when one inducible gene is regulated by one inducible system in a single cell. The Lac/Tet dual-inducible system could function in many cell lines, which was demonstrated by regulating the expression of beta-galactosidase and luciferase reporter genes in five tumor cell lines by transient transfection analysis. The feasibility of introducing a second inducible system into an already established inducible cell line was confirmed. Finally, we showed that the Lac/Tet dual-inducible system functions at translational and at functional levels in a stable cell line named 7-4-b, which contains the Ha-ras and bc1-2 inducible genes. In conclusion, this study extends the application of prokaryotic inducible systems from the regulation of a single gene to two genes and helps clarify the relationship between two genes and the effects of two genes on the cells.     

Cerebral blood flow velocity and vasomotor reactivity before and after shunting surgery in patients with normal pressure hydrocephalus

The purpose of this study was to evaluate pre- and post-shunting haemodynamic changes and their correlation with the clinical results in normal pressure hydrocephalus (NPH). Accordingly, eleven demented patients with clinical signs suggestive of NPH received examinations of cerebral blood flow velocity (BFV) and vasomotor reactivity (VMR) by transcranial Doppler sonography with carbogen testing before and after shunt treatment. Computerized tomography (CT), clinical assessment and neuropsychological grading were performed prior to and at 3 months following surgery. A control group consisting of 10 patients was included to establish baseline data. The pre-operative CBF studies in the anterior cerebral artery (ACA) and the middle cerebral artery (MCA) revealed the NPH patients did not have significant decreases of BFVs, but had significant decreases of carbogen VMR (P < 0.05). After shunting, there were no significant changes of the BFVs as compared with the pre-shunting data. The post-shunting VMR of the ACA was significantly higher than the pre-shunting one (p < 0.05), but there was no variation in that of the MCA. Both the values of post-shunting VMR in ACA and the post-shunting increase in VMR in MCA of the 7 shunt-responsive patients who improved mentally and in other symptoms were significantly higher than those of patients without improvement (p < 0.05). In addition, the five patients with gait improvement showed significantly higher values of post-shunting VMR of ACA and the post-shunting increase of VMR for both ACA and MCA when compared with those patients without gait improvement (p < 0.05, respectively). Our study supports the view that patients with NPH had various degrees of impaired VMR in both the ACA and the MCA, but showed insignificant reduction in BFVs, indicating a compensatory mechanism of CBF over time to accommodate the subnormal state of cerebral perfusion pressure. Shunt placement would improve the VMR in responsive patients. Postoperatively, an increase of VMR tends to accompany improvement of the functional state: that in the MCA alone is associated with symptomatic improvement in mental function and that increase in VMR in both the ACA and the MCA with improvement in gait, respectively.     

Non-response to maprotiline caused by ultra-rapid metabolism that is different from CYP2D6?

In this historical prospective study using sera stored for 22 years, we investigated the effect of HTLV-I infection on survival in a population of leprosy patients in the Democratic Republic of the Congo (formerly Zaire). We also determined the distribution of HTLV-I by subpopulation, age, and gender. Stored sera taken from a population of leprosy patients and controls in 1969 were tested for HTLV-I. Follow-up survival data on these patients were obtained in 1991. The sera collected in 1969 from 520 individuals was used to determine the prevalence of HTLV-I. Included in this number were 328 patients resident in the sanatorium. Survival and other data were available for 327 of these. A multivariate survival analysis using a logistic regression model was performed to evaluate the influence of HTLV-I status, age, type of leprosy, gender, duration of hospitalization, and ethnic group on survival. The overall prevalence of HTLV-I among the 520 individuals in the prevalence study was 34%, with 37.4% in the leprosy group and 25.2% in the control group (p < 0.01). Multivariate analysis using logistic regression showed that females of the Mongo and Ngombe ethnic group taken together were significantly more likely to be infected than the other groups (OR = 3.67, 95% CI: 2.14 to 6.30). A comparison of the death rates directly standardized for age and sex showed that the rate was significantly higher for HTLV-I positive (5.5/100 person-years of observation) compared with HTLV-I negative (3.6/100 person-years of observation). A survival analysis using the Cox model showed a risk ratio of 1.4 (CI: 1.04 to 1.89) for those infected with HTLV-I. An increase in the death rate was associated with HTLV-I infection in leprosy inpatients. The decreased survival associated with HTLV-I infection may result from an increased susceptibility to a variety of diseases.     

High-dose busulfan, melphalan, thiotepa and peripheral blood stem cell infusion for the treatment of metastatic breast cancer

The purpose of this study was to determine the outcome of patients with metastatic breast cancer treated with high-dose busulfan (Bu), melphalan (Mel) and thiotepa (TT) followed by peripheral blood stem cell (PBSC) infusion. Fifty-one patients with chemotherapy refractory (n = 32) or responsive (n = 19) metastatic breast cancer received Bu (12 mg/kg), Mel (100 mg/m2) and TT (500 mg/m2) followed by PBSC collected after chemotherapy and growth factor (n = 43) or growth factor alone (n = 8). The 100 day treatment-related mortality was 8% including one death from cytomegalovirus pneumonia, one from aspiration pneumonia and two from regimen-related toxicity (RRT). Seven of 28 refractory (25%) and 5/7 (71%) responsive patients with evaluable disease achieved a complete response of all measurable disease or all soft tissue disease with at least improvement in bone lesions (PR*). Fifteen of 51 patients (29%) are alive and progression-free a median of 423 days (range 353-934) after treatment, 5/32 (16%) with refractory disease and 10/19 (53%) with responsive disease. The probabilities of progression-free survival (PFS) at 1.5 years for the patients with refractory (n = 32) and responsive (n = 19) disease were 0.24 and 0.53, respectively. These preliminary data suggest that high-dose Bu/Mel/TT has significant activity in patients with advanced breast cancer and may be superior to some previously published regimens.     

扭摆式微硅隧道加速度传感器

本文介绍了扭摆式隧道加速度传感器的结构、工作原理、简化的力学模型,设计了版图、工艺流程及关键工艺,并制备出样品,最后给出测试电路和结果。     

Cloning and characterization of plastid ribosomal protein S16 gene from potato (Solanum tuberosum L. cv Désirée)

STUDY OBJECTIVE: To determine whether an intensive educational campaign of emergency department personnel on the organ donor and procurement process would result in both increased organ donor referrals and organs procured. METHODS: A retrospective review of the performance of an urban teaching ED in identifying and referring potential organ donor candidates was performed. Subsequently an intensive educational campaign of all ED staff, in conjunction with the Regional Organ Procurement Agency of Southern California (ROPA), was initiated. Physicians and nurses were educated about the procurement process, and a ROPA representative was on call 24 hours a day to assist in this process. The need for aggressive resuscitation and vital sign maintenance in potential donors as a strategy to promote organ recovery was emphasized. Reeducation by ROPA occurred every 2 to 3 months. The identification and referral rates were then retrospectively reviewed to evaluate any improvement. RESULTS: In 1994 the initial referral rate of potential organ donors from the ED was 30% (3 of 10) resulting in no organs procured. After the intervention the referral rate increased to 100% (25 of 25) in 1995 (P < .0001). The number of actual donors procured was 0 in 1994, 5 in 1995, and 9 in 1996. The increased ED referrals resulted in 14 and 32 organs procured in 1995 and 1996, respectively. CONCLUSION: Emergency physicians are in a unique position as first caregivers to interact with both potential donors and their families. With intensive education of ED staff, proper identification and referral, as well as timely intervention by organ procurement representatives, the consent and donation rate of organs for transplantation can be increased and maintained.     

The absence and application of stable carbon isotopic fractionation during the reductive dechlorination of polychlorinated biphenyls

A bacterial enrichment culture (specific to doubly flanked chlorine removal) reductively dechlorinated 2,3,4,5-tetrachlorobiphenyl (2,3,4,5-CB) to 2,3,5-trichlorobiphenyl (2,3,5-CB) in aqueous media. Approximately 90% conversion to 2,3,5-CB occurred after 90 days, with no other products formed. The delta13C values of 2,3,4,5-CB and 2,3,5-CB were relatively constant over the course of the reaction, indicating a very small or no isotope effect. In addition, compound-specific delta13C analysis performed for every congener in three different lots of Aroclor 1268 showed an intrinsic isotopic trend of decreasing 13C abundance with increasing chlorine content, similar to observations in other commercial mixtures of polychlorinated biphenyls (PCBs). The results of this laboratory study suggest that microbial reductive dechlorination of PCBs in contaminated sediments will create congeners with more depleted delta13C values than native PCBs of similar chlorination. Such information may provide additional evidence for the occurrence of this process and aid in further understanding the biogeochemistry of these compounds.     

Dual-energy x-ray absorptiometry of raloxifene effects on the lumbar vertebrae and femora of ovariectomized rats

A new potential therapeutic agent for postmenopausal osteoporosis, raloxifene, previously known as keoxifene, was evaluated by x-ray densitometry and more traditional techniques in quantitating the short-term (4-5 weeks) effects of ovariectomy on bones from 6-month-old rats. A Hologic QDR 1000/W and, to a limited extent, a Lunar DPXL, was used to quantitate ovariectomy, estrogen replacement, and raloxifene effects on vertebrae, femora, and tibiae. Both instruments performed well with precisions of 1.6% (Hologic) and 0.9% (Lunar) for anesthetized rats, which improved to 0.4% (Hologic) and 0.5% (Lunar) when the same rats were frozen. The lumbar vertebrae L1-4 showed a 12% decrease in bone mineral density 4 weeks after ovariectomy, compared with a 9% decrease for femora. Tibiae were also examined, but edge-detection problems prevented reproducible analysis of this site in vivo. The decrease in bone mineral density postovariectomy, especially for femora, was found to include both an increase in the projected area and a slight but not significant decrease in the bone mineral content of L1-4 and femora. These changes in density parameters of femora were supported by a decrease in dry weight and volume and a marginal increase in the second moment of inertia I for the identical femora examined ex vivo. Examination of individual lumbar vertebrae L1-5 suggested that the bone mineral density of L3 changes most dramatically in response to ovariectomy, but present techniques lack the spatial resolution and precision to quantitate bone changes reliably in individual vertebrae. 17 beta-Estradiol administered at 100 micrograms/kg/day subcutaneously inhibited ovariectomy effects on L1-4 bone mineral density, femoral moment of inertia, dry weight, and volume and to a lesser extent, femoral bone mineral density. A nonsteroidal compound, raloxifene HCl, at 1 mg/kg/day per os, had bone effects and effects on body weight that were largely indistinguishable from those of 17 beta-estradiol; however, raloxifene did not produce the uterotrophic effects observed with estrogen. The half-maximal efficacious dose of raloxifene on L1-4 bone mineral density was between 0.1 and 1.0 mg/kg/day per os. These data show that dual-energy x-ray absorptiometry compares favorably with traditional methods in quantitating bone changes caused by ovariectomy in small rodents, that L1-4 is a more sensitive region than whole femora in evaluating the effect of estrogen deficiency on bone loss, and the raloxifene may have promise as a treatment for conditions characterized by excessive bone loss after ovariectomy.     

Intraosseous pressure in Kienb?ck's disease

Intraosseous pressure was measured in the lunate of 10 patients undergoing surgery for Kienb?ck's disease. Using the styloid process of the radius and the capitate as reference points of bone pressure, I found a stasis rather than arterial insufficiency in the lunate. This pattern was observed regardless of the stage of the disease according to St?hl and regardless of the finding by technetium TC 99m-methylene diphosphonate scintigraphy. The significance of this finding is still unknown but questions the current concept that Kienb?ck's disease is caused by arterial insufficiency, because these results imply an interruption of the blood flow as a result of venous congestion.