A scanning micropipette molecule microscope.

A movable quartz micropipette, whose tip is sealed with a polymer plug, is used as a liquid-vacuum interface to a mass spectrometer. A light microscope allows observation of, and positioning of, the micropipette tip on the surface of a sample mounted in a perfusion chamber. This forms the basis of an instrument which enables one to study, in vitro, the localization of transepithelial transport of water and other molecules. Some preliminary results from the use of this instrument are presented.

Inhibition of human brain tumor cell growth by the anti-inflammatory drug, flurbiprofen.

Despite many efforts to alter the relentlessly aggressive progression of tumors of neural origin, individuals bearing these tumors exhibit poor prognosis for long-term survival. In an attempt to find an effective treatment, we examined the efficacy of the non-steroidal anti-inflammatory drug, flurbiprofen, to suppress the growth of tumor cell lines derived from medulloblastoma and glioblastoma multiforme. Results from cell proliferation assays have revealed that flurbiprofen effectively inhibits the growth of various tumor cells in a dose-dependent manner and causes a noticeable change in the progression of cells through cell cycle stages. Treatment of tumor cells with flurbiprofen reduced the number of cells in G1 and G2, and significantly increased their numbers in S phase, suggesting that, flurbiprofen accelerates G1/S entry, and/or delays cell exit from S to G2/M stages. Results from RNase protection assay and Western blot analysis showed that while treatment of cells with flurbiprofen causes a minor change in the RNA level of different cyclins, there is a significant decrease in the level of cyclin B protein upon flurbiprofen treatment. Examination of tumor suppressors by RNase protection technique showed a subtle increase in the levels of several tumor suppressors upon flurbiprofen treatment. Interestingly, at the protein level, p53 tumor suppressor was substantially increased upon flurbiprofen treatment, yet the level of p21, a downstream target for p53 remained unchanged. Curiously, treatment of the cells with flurbiprofen enhanced the level of COX-2 expression. Results from co-immunoprecipitation showed association of COX-2 with p53 in tumor cells. These observations suggest that the interaction of COX-2 with p53 may cause p21-independent suppression of tumor cell growth upon flurbiprofen treatment.

Time responses of cancellous and cortical bones to sciatic neurectomy in growing female rats.

Effects of unilateral sciatic neurectomy on the responses of both cancellous and cortical bones were studied in growing female rats at 0, 1, 4, 8, and 12 weeks after operation. Using double-fluorescent labeling techniques, histomorphometric analyses were performed on longitudinal sections of proximal tibial metaphyseal secondary spongiosa (PTM) and on cross sections of tibial shaft (TX). In PTM, sciatic neurectomy not only inhibited the age-related bone gain, but also reduced the trabecular bone mass by 46%, which was accompanied by decreases in trabecular number, thickness, and node to node density, and an increase in trabecular separation and free end to free end density. The bone loss occurred mainly between 1 and 4 weeks after operation. A sharp increase in bone formation indices was observed during the first week after nerve section. However, these endpoints quickly dropped to levels lower than those of sham-operated controls at 4 weeks, and were not different from the control levels at 8 weeks after operation. Eroded surface increased progressively after sciatic neurectomy during the 12 weeks experimental period. In TX, sciatic neurectomy inhibited the age-related increase in total tissue area that maintained it at the basal control level. However, the cortical bone area in neurectomized legs was lower than that in sham-operated controls. Sciatic neurectomy also stimulated the bone formation indices on both periosteal and endocortical surfaces during the first week after operation. These endpoints declined sharply between 1 and 4 weeks and then maintained at control levels between 8 and 12 weeks post surgery. Endocortical eroded surface increased 1 week after neurectomy, reached the peak at 8 weeks, and then decreased thereafter. These findings suggest that (1) sciatic neurectomy not only inhibited age-related bone gain but also induced marked bone loss in cancellous bone site and inhibited age-related bone gain in cortical bone site, which mainly resulted from the decrease in bone formation and the increase in bone resorption; (2) the changes in both cancellous and cortical bones responded to sciatic neurectomy occurred mostly within the first 4 weeks and stabilized between 8 and 12 weeks after surgical intervention. In conclusion, the unilateral sciatic neurectomized rat is a complex model in which to study osteopenia. Despite sciatic neurectomy being a simple operation, the interactions of skeletal responses to postsurgical regional acceleratory phenomenon (RAP) and disuse and adaptation changes cannot be clearly differentiated. Furthermore, the complications from growth and aging should be avoided.

The potential effects of radiographic criteria to exclude aortography in patients with blunt chest trauma. Results of a study of 32 patients with proved aortic or brachiocephalic arterial injury.

The purpose of this study was to test the effectiveness, in patients with known aortic or brachiocephalic arterial injury, of five previously published radiographic criteria for excluding aortography in patients with blunt chest trauma. These criteria were (1) normal findings on erect chest radiograph; (2) normal aortic arch and left subclavian artery; (3) normal aortic arch, descending aorta, aortopulmonary window, tracheal position, and left paraspinal interface; (4) normal right paratracheal stripe and nasogastric tube position, and (5) normal aortic arch and tracheal and nasogastric tube position. One or more of these criteria were met in 6% to 25% of patient with major thoracic arterial injury, depending on the criteria used. Interestingly, two (6%) patients had radiographs that showed no specific signs of mediastinal hemorrhage, which indicates that the chest radiograph is limited in its sensitivity to detect major thoracic arterial injury. Because of these results, we do not believe that attempts to limit aortography in patients with supine film evidence of mediastinal abnormality, based on the absence of certain signs of mediastinal hemorrhage, are warranted. Furthermore, an abnormal radiograph cannot be relied on as the sole criterion for aortography if the goal of care is to detect as close to 100% of vascular injuries as possible.

Selective, unilateral, reversible loss of behavioral responses to looming stimuli after injection of tetrodotoxin of cadmium chloride into the frog optic nerve.

Unilateral injection of tetrodotoxin or cadmium chloride into the frog optic nerve selectively eliminates behavioral responses to looming stimuli while sparing responses to prey stimuli. This behavioral loss is correlated with a loss of activity of "dimming" units in tectal layer G. These findings suggest that separate sets of retinal ganglion cell fibers carry information concerning looming stimuli and prey stimuli. The lack of activity in layer G suggests that information about looming stimuli is being conveyed by myelinated retinal ganglion cell axons. It is argued that unmyelinated fibers are not blocked by the neurotoxins because the extracellular space around the fibers is mostly inaccessible.

Evaluating the sale of a nonprofit health system to a for-profit hospital management company: the Legacy Experience.

OBJECTIVE: To introduce and develop a decision model that can be used by the leadership of nonprofit healthcare organizations to assist them in evaluating whether selling to a for-profit organization is in their community's best interest. STUDY SETTING/DATA SOURCES: A case study of the planning process and decision model that Legacy Health System used to evaluate whether to sell to a for-profit hospital management company and use the proceeds of the sale to establish a community health foundation. Data sources included financial statements of benchmark organizations, internal company records, and numerous existing studies. STUDY DESIGN: The development of the multivariate model was based on insight gathered through a review of the current literature regarding the conversion of nonprofit healthcare organizations. DATA COLLECTION/EXTRACTION METHODS: The effect that conversion from nonprofit to for-profit status would have on each variable was estimated based on assumptions drawn from the current literature and on an analysis of Legacy and for-profit hospital company data. PRINCIPAL FINDINGS: The results of the decision model calculations indicate that the sale of Legacy to a for-profit firm and the subsequent creation of a community foundation would have a negative effect on the local community. CONCLUSIONS: The use of the decision model enabled senior management and trustees to systematically address the conversion question and to conclude that continuing to operate as a nonprofit organization would provide the most benefit to the local community. The model will prove useful to organizations that decide to sell to a for-profit organization as well as those that choose to continue nonprofit operations. For those that decide to sell, the model will assist in minimizing any potential negative effect that conversion may have on the community. The model will help those who choose not to sell to develop a better understanding of the organization's value to the community.

Endogenous natriuretic factors 3: isolation and characterization of human natriuretic factors LLU-alpha, LLU-beta 1, and LLU-gamma.

A low molecular weight endogenous substance believed to be responsible for extracellular fluid homeostasis in mammals has been sought for many years. Our goal is to isolate and structurally characterize this putative "natriuretic hormone." We have developed an assay using the conscious rat to measure prolonged natriuresis (Benaksas et al (1993) Life Sciences, 52, 1045-1054), the activity originally described for this putative substance. Using this assay we have identified a number of natriuretic compounds isolated from human uremic urine. The collected urine is processed by ultrafiltration (< or = 3 kDa), gel filtration chromatography (G-25) and extraction with isopropanol and diethyl ether. The organic soluble material is then subjected to sequential high-performance liquid chromatography. We report here the initial characterization of two pure isolates (LLU-alpha and LLU-gamma) obtained by this method, and the structural elucidation of a third pure compound, LLU-beta 1, a natriuretic and previously unreported metabolite of the drug diltiazem.

Acting on mission to care for the needy.

Evangelical Health Systems (EHS), Chicago's largest provider of health care services, through long-range strategic planning continues to fulfill its religious-based mission of caring for the needy. By concentrating on the strengths and needs of the residents of the West and South Sides, EHS is developing programs and services to care for the elderly and poor. This article outlines key components of the system's strategy in these areas.

A religiously affiliated health care system in today's competitive market: a success story.

In today's competitive health care market, not-for-profit religiously affiliated health care systems face multiple challenges for survival. Their mission of providing for the effective and efficient delivery of quality health care and health-related services for the benefit of individuals, families, and society and a philosophy of maintaining a Christian emphasis in all endeavors conflicts with many of the current competitive themes in health care. The successful system profiled here is Evangelical Health Systems (EHS), a not-for-profit organization affiliated with the United Church of Christ.

1991 outlook: progress will require partnerships, improved productivity--roundtable discussion.

To address the myriad problems and challenges in the next year, the operative word will be partnership. That's according to members of Modern Healthcare's editorial advisory board in their discussion of the healthcare industry in 1991. The experts see the need for hospitals, physicians and the business community to team up to control costs, solve staffing woes and take initial steps toward healthcare reform.

Building a legacy for the future: creating an integrated health care system.

Local and national market forces of decreasing inpatient utilization and increasing costs paved the way for a major merger involving five hospitals and one home health agency in Portland, Oregon. The result: a health care delivery system that merged corporate cultures, eliminated duplicity, integrated medical and administrative staffs, realized cost containment savings, and effectively responded to market trends toward reform.

Determination of normal transverse mediastinal width and mediastinal-width to chest-width (M/C) ratio in control subjects: implications for subjects with aortic or brachiocephalic arterial injury.

We measured transverse mediastinal width and mediastinal-width to chest-width (M/C) ratio on supine films of 100 nontraumatized controls. In 95% the transverse mediastinal width was less than 7.5 cm and the M/C ratio was less than 0.38. Thus a transverse width of 7.5 cm or more or an M/C ratio of 0.38 or more can be defined as abnormal with 95% confidence. Application of these values to determine abnormality in 32 patients with proven aortic or brachiocephalic injury showed that the transverse mediastinal width was within normal limits in 41% and M/C ratio was normal in 69%. Utilizing smaller values that would identify all abnormals resulted in false positive rates in the controls of 74% and 87%, respectively. However, one or more of eight specific signs of mediastinal abnormality related to hemorrhage or pseudoaneurysm formation were present in 94% of abnormals compared to only 11% of controls. Because of extreme overlap of transverse mediastinal width and M/C ratio between normals and abnormals, precise measurement of the mediastinum cannot reliably separate the two groups. The subjective assessment of anatomic mediastinal abnormality remains a superior plain film method in determining the need for aortography.

The molecule microscope: a new instrument for biological and biomedical research.

We describe a new instrument, the molecule microscope, which reveals directly spatial variations in the rate of evaporation of molecules from surfaces by using neutral molecules instead of light or charged particles used in existing kinds of microscopes. The surface composition of the sample determines the binding energy of the evaporating molecules and, hence, the rate of evaporation, which also depends on permeability of the sample when the molecules come either from within or from the other side. We show first results obtained with our apparatus, discuss the design of an instrument now under construction with approximately 1-mum resolution, and describe briefly some more advanced versions under consideration with approximately 100-A resolution.

Use of mass spectrometer to measure CO2 and O2 fluxes in voltage-clamped epithelia.

A quadrupole mass spectrometer was coupled to an Ussing chamber in order to evaluate rates of oxidative metabolism in voltage-clamped epithelia. Well-defined mixing characteristics of the continuously perfused chamber allowed CO2 and O2 concentrations to be related to rates of CO2 efflux, JCO2, and oxygen influx, JO2. The use of a model tissue to simulate step changes in JCO2 validated the treatment, with response within a minute. Monitoring of metabolism was facilitated by use of a desk-top computer, which evaluated JCO2 at 6-s intervals. Concurrent measurements of electrical current and JCO2 were made in the toad urinary bladder in order to relate active sodium transport to metabolism; the use of amiloride to eliminate active transport and the associated metabolism then allowed evaluation of the rates of active Na transport (JNa) and suprabasal metabolism (JsbCO2), and their ratio JNa/JsbCO2. We report the ability to resolve a 5 pmol/s change in CO2 efflux or an 11 pmol/s change in O2 influx rates.

Time course of active Na transport and oxidative metabolism following transepithelial potential perturbation in toad urinary bladder.

The use of an Ussing chamber with well-defined mixing characteristics coupled to a mass spectrometer permits the concurrent evaluation of transepithelial current and oxidative metabolism with improved temporal resolution. The time-course of the amiloride-sensitive current Ia and the rate of suprabasal CO2 production JsbCO2 were observed in 10 toad urinary bladders at short-circuit and after clamping delta psi at 100 mV, serosa positive. Following perturbation of delta psi (0 leads to 100 mV), Ia declined sharply within 1/2 min, remaining near constant approximately 15 min, and then increased slightly. JsbCO2 declined more gradually, remained near constant at approximately 4-7 min, and then declined further. Detailed analysis revealed an early quasi-steady state with near constancy of JsbCO2 starting at 2.9 +/- 1.1 (SD) min and lasting 4.7 +/- 1.8 (SD) min, followed by relaxation to a later steady state at about 15 min. During the early quasi-steady state, Ia was also nearly constant. Considering that in steady states Ia/F approximately or equal to JaNa, the rate of transepithelial active Na transport, during the early quasi-steady state mean values +/- SE of JaNa, JsbCO2 and (JaNa/JsbCO2) were, respectively, 29.9 +/- 1.7%, 59.4 +/- 3.2%, and 56.4 +/- 5.7% of values at short-circuit. Corresponding values during the late steady state were 41.4 +/- 6.0%, 38.2 +/- 6.1%, and 111.3 +/- 8.6%. Thus the flow ratio JaNa/JsbCO2 was depressed significantly during the early quasi-steady state, but returned later to the original value. The results of measurements of Ia and JsbO2 in three hemibladders were qualitatively similar. In terms of a phenomenological "black-box" treatment the findings are consistent with earlier studies indicating incomplete coupling between transport and metabolism. Further studies will be required to clarify the molecular basis for these observations.