Faculty Bibliography

BACKGROUND: Insulin attenuates serotonin-induced Ca2+ influx, the intracellular Ca2+ transient, and contraction of cultured vascular smooth muscle cells from dog femoral artery. These studies were designed to test whether insulin-induced glucose transport was an early event leading to the inhibitory effects of insulin on Ca2+ influx, intracellular Ca2+ concentration, and contraction in these cells. METHODS AND RESULTS: Insulin 1 nmol/L stimulated the 30-minute uptake of [3H]2-deoxyglucose in these cells via a phloridzin-inhibitable mechanism. Contraction of individual cells was measured by photomicroscopy, intracellular Ca2+ concentration was monitored by measuring fura 2 fluorescence by use of Ca(2+)-sensitive excitation wavelengths, and Ca2+ influx was estimated by the rate of Mn2+ quenching of intracellular fura 2 fluorescence when excited at a Ca(2+)-insensitive wave-length. In the presence of 5 mmol/L glucose, preincubation of cells for 30 minutes with 1 nmol/L insulin inhibited 10(-5) mol/L serotonin-induced contraction of individual cells by 62% (P < .01) and decreased the serotonin-stimulated component of Mn2+ influx by 78% (P < .05). Removing glucose from the preincubation medium or adding 1 mmol/L phloridzin completely eliminated these effects of insulin. Insulin lowered the serotonin-induced intracellular Ca2+ peak by 37% (P < .05), and phloridzin blocked this effect of insulin. When glucose uptake was increased to the insulin-stimulated level by preincubation of the cells for 30 minutes with 25 mmol/L glucose in the absence of insulin, serotonin failed to stimulate Mn2+ influx, the serotonin-induced Ca2+ peak was decreased by 46% (P < .05), serotonin-induced contraction was inhibited by 60% (P < .01), and addition of insulin did not further inhibit contraction. CONCLUSIONS: Since the effects of insulin on serotonin-stimulated Ca2+ transport, intracellular Ca2+ concentration, and contraction were dependent on glucose transport and were duplicated when glucose transport was stimulated by high extracellular glucose concentration rather than insulin per se, it is concluded that insulin-stimulated glucose transport is an early event that leads to decreased Ca2+ influx and contraction in vascular smooth muscle.

A dialysis method was developed for measuring free Zn2+ concentration in plasma and milk for determination of the electrochemical gradient for Zn2+ across the mammary gland. This method used the zinc content of casein after dialysis as the metal ion indicator because zinc in the calcium phosphate-citrate complex inside casein micelles is dependent on the free Zn2+ concentration of an associated aqueous phase. Zinc, calcium and magnesium distribution in milk confirmed the high zinc binding by bovine casein. Zinc in the casein, dialyzed against standards or unknowns, was measured by atomic absorption spectrophotometry. Average free Zn2+ concentrations measured by the dialysis method in plasma and milk of five cows were 0.141 and 0.331 nmol/L, respectively. The equilibrium potential of Zn2+ ions across the mammary epithelium, calculated from free Zn2+ concentrations in blood and milk measured by the dialysis method, was -11.4 mV, consistent with the mammary electrical potentials noted in previous studies. Therefore, no electrochemical gradient for zinc between the two compartments was apparent, and it is not necessary to invoke an active transport mechanism in the mammary gland to explain the higher zinc concentration in milk than in plasma of most species.

PURPOSE: To examine the effects of lomustine (CCNU), a commonly used nitrosourea, and craniospinal radiation therapy on the subsequent development of restrictive lung disease (RLD) following treatment for malignant brain tumors. PATIENTS AND METHODS: Pulmonary function testing with measurement of lung volume, spirometry, and diffusion capacity was performed in 28 patients who had received CCNU and/or radiation therapy as treatment for a malignant brain tumor. The median age at the time of treatment was 11.4 years (range, 3.9 to 36.7) and radiation therapy was completed 6 months to 11.6 years (median, 2.6 years) before testing. Patients were divided into four groups based on prior therapy. Group 1 received involved-field irradiation and a CCNU-containing chemotherapy regimen (n = 7); group 2, craniospinal irradiation with a boost to the primary tumor site and a CCNU-containing chemotherapy regimen (n = 6); group 3, craniospinal irradiation with a boost to the primary tumor site and a non-CCNU-containing chemotherapy regimen (n = 7); and group 4, craniospinal irradiation with a boost to the primary tumor site without chemotherapy (n = 8). RESULTS: Fourteen patients (50%) had findings consistent with RLD. One of seven patients (14.3%) who received CCNU without spinal irradiation had RLD, whereas 13 of 21 (61.9%) who received spinal irradiation with or without CCNU had RLD (P = .038), including four of eight patients treated with craniospinal irradiation alone. Logistic regression analysis showed that only spinal irradiation was a significant predictor for RLD. Patients who received spinal irradiation were 4.3 times more likely to have RLD than those who did not receive spinal irradiation. CONCLUSION: Spinal irradiation may be a risk factor for the development of RLD

PURPOSE: To describe the biologic and clinical features of children with primitive neuroectodermal tumors (PNETs) arising in the pineal region (pineoblastomas) and evaluate prospectively the efficacy of radiation therapy (RT) and/or chemotherapy. PATIENTS AND METHODS: Between 1986 and 1992, 25 children with PNETs of the pineal region were treated as part of a Childrens Cancer Group study. Eight infants less than 18 months of age were nonrandomly treated with eight-drugs-in-1-day chemotherapy without RT. The remaining 17 patients were treated with craniospinal RT and randomized to receive either vincristine, lomustine (CCNU), and prednisone or the eight-drugs-in-1-day regimen. RESULTS: Of 24 completely staged patients, 20 (83%) had localized disease at diagnosis. All infants developed progressive disease a median of 4 months from the start of treatment. Of the 17 older patients treated with RT and chemotherapy, the Kaplan-Meier estimate of progression-free survival (PFS) at 3 years is 61% +/- 13%. This is superior to the PFS of children with other supratentorial PNETs (P = .026). Following RT, 12 of 17 patients (70.6%) had a residual pineal region mass, which persisted for as long as 5 years before resolving; only four subsequently developed progressive disease. CONCLUSION: (1) Eight-in-1 chemotherapy without RT appears to be ineffective therapy for young children with PNETs of the pineal region. (2) For children more than 18 months of age at diagnosis treated with craniospinal RT and chemotherapy, the PFS is superior to that of children with other supratentorial PNETs. (3) A residual enhancing mass following RT is not predictive of treatment failure

PURPOSE: Cyclophosphamide is commonly used in the treatment of children with malignant brain tumors. The purpose of this study was to develop a multicycle, high-dose intensity cyclophosphamide regimen with granulocyte-macrophage colony-stimulating factor (GM-CSF) and to assess its activity against malignant glioma and primitive neuroectodermal tumor (PNET). METHODS: Twenty-three patients with brain tumors, including 15 with malignant glioma and six with PNET, were enrolled. Cyclophosphamide (1.8-2.25 g/m2/day for 2 days i.v.; total dose 3.6-4.5 g/m2) was administered and was followed by recombinant human GM-CSF (5 micrograms/kg/day s.c.) on days 3-11 or until the absolute granulocyte count reached 1.5 x 10(9)/L. RESULTS: With a total of 83 cycles administered, the mean dose intensity of cyclophosphamide ranged from 1.5 g/m2/week through cycle 2 (22 patients) to 0.8 g/m2/week through cycle 8 (two patients). No activity was seen against malignant glioma, and five of six patients with PNET had partial responses. The mean duration of a neutrophil count of < 0.5 x 10(9)/L was only 8 days; the platelet recovery was substantially longer. Fever during neutropenia occurred in 54 of 83 cycles. One patient died from transfusion-related graft-versus-host disease. CONCLUSIONS: A cyclophosphamide regimen equal to twice the dose intensity of that used in conventional therapy was administered. The regimen was active against PNET but inactive against malignant glioma

PURPOSE/OBJECTIVE:This study was undertaken to evaluate the radiographic response to two cycles of chemotherapy prior to irradiation in newly diagnosed children with high-grade astrocytomas. PATIENTS AND METHODS/METHODS:One hundred and thirty children less than 21 years of age with newly-diagnosed high-grade astrocytoma were treated with the 'eight-drugs-in-one-day' chemotherapy regimen as part of a phase III multi-institutional Childrens Cancer Group (CCG) trial. Computerized Tomographic (CT) or Magnetic Resonance Image (MRI) scans, obtained after two cycles of chemotherapy had been administered, were compared with post-operative scans to determine treatment response. Scans were evaluated by institutional radiologists, and were reviewed centrally by a single neuroradiologist. RESULTS:Of 79 patients with evaluable post-operative residual tumor on CT or MRI scans, 26 (33%) were determined on institutional evaluation to have had an objective response. However, central review of scans documented responses on only 14/79 (18%). A significantly higher response rate on central review was observed for those children 36 months of age or less at study entry than for older children (33% v 11%; p < 0.001). However, a higher disease progression rate was also observed for those children 36 months of age or less than for older children (21% v 2.6%; p < 0.001). CONCLUSION/CONCLUSIONS:In this study, the largest yet reported in newly-diagnosed children with high-grade astrocytomas, the chemotherapy regimen has activity in younger children. The differences in response rates reported by institutional and central review highlight the difficulties inherent in assessing response to brain tumor therapy. However, the study does demonstrate the consistent ability of radiologists to identify disease progression within the institutional and central reviews.

BACKGROUND: Insulin in physiological concentrations attenuates the agonist-induced intracellular Ca2+ ([Ca2+]i) transient and inhibits contraction in individual nonproliferated cultured canine femoral artery vascular smooth muscle cells (VSMCs). In the present study, we wished to define the effects of insulin on individual components of Ca2+ transport in vascular smooth muscle. METHODS AND RESULTS: Insulin (40 microU/mL) attenuated the 5-hydroxytryptamine (5-HT, serotonin; 10(-5) mol/L)-induced [Ca2+]i transient (measured by fura 2 fluorescence) in primary confluent canine femoral artery VSMCs in the presence of extracellular Ca2+. In Ca(2+)-free media, the 5-HT-induced [Ca2+]i transient was reduced by 42% and was not affected by insulin. This finding suggested that insulin inhibits 5-HT-induced Ca2+ influx but does not affect sarcolemmal Ca2+ efflux or Ca2+ release from intracellular stores. In support of those conclusions, we found that insulin inhibited the 5-HT-induced component of Mn2+ (a Ca2+ surrogate) influx (measured by fura 2 fluorescence quenching at the Ca2+ isosbestic excitation wavelength). In addition, 5-HT stimulated the rates of 45Ca2+ efflux from intact cells (a measure of sarcolemmal Ca2+ efflux) and from saponin-permeabilized cells (a measure of Ca2+ release from intracellular stores), but insulin did not affect these rates of 45Ca2+ efflux. CONCLUSIONS: We conclude that a physiological insulin concentration attenuates the 5-HT-induced [Ca2+]i transient in confluent primary cultured canine femoral artery VSMCs by inhibiting the 5-HT-induced component of Ca2+ influx but not by affecting sarcolemmal Ca2+ efflux or Ca2+ release from intracellular stores.

The partitioning of calcium in human milk was studied experimentally and compared with the distribution of calcium among the compartments of bovine milk. Care was taken to handle the milk samples to produce minimal disturbance of the milk fat globule membranes and the CO2 content. About 15% of the milk calcium was associated with casein in human milk; the remainder was in the aqueous phase. The equilibrium between ionized calcium and calcium complexes with smaller anions was examined. Ionized calcium varied between 2.3 and 4.0 mM across individuals (mean 3.0 +/- .1 mM) at 90 d of lactation. The remainder of the calcium in the aqueous phase was associated with citrate and phosphate. Changes in total calcium in human milk during lactogenesis and late lactation reflected changes in citrate and casein rather than alterations in ionized calcium.