Effects of growth hormone, insulin-like growth factor-I, and cortisol on periparturient antibody response profiles of dairy cattle

The objectives of this study were to determine hormone and antibody response profiles from the prepartum period to peak lactation, and evaluate potential immunomodulatory effects of the classic endocrine hormones, growth hormone (GH), insulin-like growth factor-I (IGF-I) and cortisol. Specifically, 33 Holstein cows were immunized with ovalbumin (OVA) and Escherichia coli J5 at weeks -8 and -3 prior to parturition. At parturition (week 0), cows received an additional immunization of OVA. Blood was collected at weeks -8, -3, 0, 3 and 6 relative to parturition and various samples were used to determine plasma hormone concentration, serum immunoglobulin (Ig), and specific antibody response to OVA and E. coli. Colostrum and milk samples were also collected post-parturition to monitor local immunoglobulin and antibody responses. Results indicated that not all periparturient cows exhibited depressed immune response, and that antibody response to OVA could be used to partition cows into 3 groups recognizing animals with sustained measurable antibody response before and after parturition (Group 1), animals which responded poorly to immunization at parturition (Group 2), and animals which did not respond to immunizations at week -3 or parturition (Group 3). Cows with the highest antibody response to OVA (Group 1) also tended (P < or = 0.10) to have the highest response to E. coli J5 at parturition and had the lowest incidence of disease, particularly mastitis. Antibody response to OVA measured in milk tended to be higher in Group 1 cows, particularly at week 0 (P < or = 0.06) compared to cows of Group 3. IGF-I was higher (P < or = 0.05) in cows of Group 1 than Group 3 at peak lactation (week 6).     

A comparison of thromboelastography with heparinase or protamine sulfate added in vitro during heparinized cardiopulmonary bypass

Thromboelastography (TEG) has been used after cardiopulmonary bypass (CPB) to diagnose excessive postoperative hemorrhage. Conventional TEG during CPB is not possible due to the sensitivity of the TEG to even small amounts of heparin, which produces a nondiagnostic tracing. The purpose of this study was to compare heparin neutralization using heparinase or protamine in TEG blood samples obtained during CPB. TEG testing was performed on 48 patients before, during and after CPB. Tissue plasminogen activator activity and antigen were measured on a subset of 32 patients. We found: 1) heparinase neutralized at least 10 IU/ml heparin while 1.6 ug/ml protamine neutralized up to 7 IU/ml heparin, 2) in samples with complete heparin neutralization by both methods, there was no significant difference in the R values, 3) while there was good correlation for other TEG parameters between heparinase and protamine treated samples, heparinase treatment produced shorter K values and higher angle, MA and A60, 4) while fibrinolysis was detected using both methods, heparinase treatment suppressed fibrinolysis in the TEG in both samples from patients and after in vitro addition of tissue plasminogen activator, 5) TEG was not a sensitive indicator of t-PA activity, detecting only 21% of samples with increased t-PA activity during bypass, and 5) heparinase was at least 100 times more expensive than protamine. We conclude that while both heparinase and protamine can be used to neutralize heparin in TEG samples obtained during CPB, protamine neutralization is more sensitive to fibrinolysis and less expensive, but the protamine dose must be carefully selected to match the heparin level used at individual institutions.     

Maturation of the posterolateral spinal fusion and its effect on load-sharing of spinal instrumentation. An in vivo sheep model

We investigated the temporal relationship among the biomechanical, radiographic, and histological properties of a posterolateral spinal fusion mass to elucidate the changes in load-sharing of the spinal instrumentation and that of the fusion mass throughout the healing process. Destabilization of the posterior spinal column and transpedicular screw fixation at the segments between the third and fourth and the fifth and sixth lumbar vertebrae was performed in twenty-four sheep. A posterolateral spinal arthrodesis with use of autologous corticocancellous bone graft was done randomly at one of the two segments; the other segment (without bone graft) served as the instrumented control. Six animals each were killed at four, eight, twelve, and sixteen weeks postoperatively. Biomechanical testing showed that the posterolateral fusion mass had increased mechanical stiffness after the fourth week. The strain on the hardware, measured with use of rods instrumented with strain-gauges, decreased significantly (p < 0.01) beginning at eight weeks. Radiographically, three independent observations of each of the six animals at each time-period showed that, although all of the fusion masses were considered solid unions at sixteen weeks, bridging of trabecular bone was noted during only ten of eighteen observations at twelve weeks, three of eighteen observations at eight weeks, and none of eighteen observations at four weeks. Computerized tomography and histomorphometric analyses demonstrated that mineralization in the fusion mass increased in a linear fashion even after eight weeks. Histologically, the fusion mass consisted predominantly of woven bone at eight weeks; thereafter, it was gradually trabeculated. CLINICAL RELEVANCE: We found a great discrepancy between biomechanical stability and histological maturation of the posterolateral fusion mass. The biomechanical properties of a stable spinal fusion preceded the radiographic appearance of a solid fusion by at least eight weeks, suggesting that immature woven bone provided substantial stiffness to the fusion mass. The spinal instrumentation was subjected predominantly to bending stress rather than to axial stress, and the load-sharing of the spinal instrumentation decreased concurrently with the development of the spinal fusion.     

Acute mast cell leukemia disclosed by vasomotor flushing

INTRODUCTION: Acute mast cell leukemia is a rare and severe disease. We report herein a case associated with a flush syndrome. CASE REPORT: A 44-year-old man, presented with a flush of face and trunk. Bone marrow was infiltrated with immature mast cells. In spite of chemotherapy and bone marrow transplantation the patient deceased. DISCUSSION: Pheochromocytoma, carcinoid tumor, and mastocytosis are associated with a flush syndrome. In our patient the diagnosis was an acute mast cell leukemia. Acute mast cell leukemia can follow systemic mastocytosis or occur de novo. This disease is of poor prognosis. No treatment is available.     

Differential divalent cation requirements uncouple the assembly and catalytic reactions of human immunodeficiency virus type 1 integrase

Previous in vitro analyses have shown that the human immunodeficiency virus type 1 (HIV-1) integrase uses either manganese or magnesium to assemble as a stable complex on the donor substrate and to catalyze strand transfer. We now demonstrate that subsequent to assembly, catalysis of both 3' end processing and strand transfer requires a divalent cation cofactor and that the divalent cation requirements for assembly and catalysis can be functionally distinguished based on the ability to utilize calcium and cobalt, respectively. The different divalent cation requirements manifest by these processes are exploited to uncouple assembly and catalysis, thus staging the reaction. Staged 3' end processing and strand transfer assays are then used in conjunction with exonuclease III protection analysis to investigate the effects of integrase inhibitors on each step in the reaction. Analysis of a series of related inhibitors demonstrates that these types of compounds affect assembly and not either catalytic process, therefore reconciling the apparent disparate results obtained for such inhibitors in assays using isolated preintegration complexes. These studies provide evidence for a distinct role of the divalent cation cofactor in assembly and catalysis and have implications for both the identification and characterization of integrase inhibitors.     

Catheter infection in intensive care: influence of systematic replacement of central venous catheters on a guide wire every 4 days

OBJECTIVES: Determine whether systematic replacement of central venous catheters on a guide wire every 4 days leads to a lower rate of catheter infection. METHODS: We conducted a prospective randomized study comparing systematic catheter replacement with the standard procedure where one-way catheters are left in situ as needed. RESULTS: The study included 150 patients and 170 catheters. Catheter replacement on a guide wire was performed 169 times. The rate of infection was not significantly different between the group of patients with replacement (3.7 for 1000 catheter days) and the group with systematic replacement (5.4 for 1000 catheter days). No mechanical complications occurred during the replacement procedures on guide wires. CONCLUSION: We do not recommend systematically replacing catheters on a guide wire for the prevention of catheter infection. This procedure may however be indicated in case of suspected catheter infection as no mechanical of infectious complications occurred.     

Atorvastatin: an effective lipid-modifying agent in familial hypercholesterolemia

Hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase inhibitors are the drugs of choice in heterozygous familial hypercholesterolemia (FH), which has a high risk of ischemic heart disease. An open-label study was conducted to test the efficacy and safety of atorvastatin, a new synthetic HMG-CoA reductase inhibitor in proven FH. After a 4-week placebo phase, 22 subjects were randomized to either 80 mg atorvastatin at night (n = 11) or 40 mg twice a day for 6 weeks. The two dosage groups were well matched and had no difference in lipoprotein responses. After 6 weeks, the LDL cholesterol concentration was reduced by 57%, from 8.16 +/- 1.15 to 3.53 +/- 0.99 mmol/L (P < .001). The total cholesterol concentration decreased from 9.90 +/- 1.32 to 5.43 mmol/L (P < .001). HDL cholesterol concentration increased from 1.19 +/- 0.31 to 1.49 +/- 0.43 mmol/L (P < .001). Triglyceride concentrations decreased from 1.34 +/- 0.66 to 0.88 +/- 0.36 mmol/L (P < .01). Three subjects had single, transient increases of serum transaminase of up to twice the upper limit of normal. Apolipoprotein B concentration decreased significantly by 42%. Changes in apolipoproteins AI and (a) were not statistically significant. Nondenaturing gradient gel electrophoresis revealed increases in the size of smaller LDL particles in four subjects. Plasma fibrinogen concentration increased by 44%. The drug was well tolerated. One subject withdrew for personal reasons. Atorvastatin is a powerful and safe lipid-modifying agent for LDL cholesterol; it also modifies HDL cholesterol and triglyceride concentrations, and may suffice as a single agent for many subjects with heterozygous FH.     

Frequency and impact of SMBG on glycemic control in patients with NIDDM in an urban teaching hospital clinic

Alternative therapy for refractory leukemic patients is being increasingly adopted. Circumvention of multidrug resistance represents a strategy that has been taken into account when conventional chemotherapy failed. In this work a group of 15 refractory, heavily pretreated, patients was enrolled in a circumvention protocol including etoposide (ETO) and cyclosporin A (CSA). All patients received etoposide prior to this schedule. Toxicity to circumvention protocol was acceptable and only one serious side-effect was observed. Two hematological clinical responses were seen, both of which were positive to P-glycoprotein immunostaining and exhibited in vitro modulation by CSA in cultures using the thymidine incorporation assay. Three out of four patients negative for P-glycoprotein achieved a minor response. Three out of six clinical failures were also negative for Pgp immunostaining one of which exhibited sinergistic effect between ETO and CSA. Our study suggests that hematological response to ETO and CSA association can be obtained in intensely pretreated leukemic patients. Several factors may affect the response such as clinical status before this therapy. Additionally, it also suggests that not all CSA effects on the combination ETO-CSA can be attributed to Pgp modulation.     

Induction of human breast cancer cell apoptosis from G2/M preceded by stimulation into the cell cycle by Z-1,1-dichloro-2,3-diphenylcyclopropane

We have shown previously that Z-1,1-dichloro-2,3-diphenylcyclopropane (a.k.a. Analog II, A(II)) inhibits human breast cancer cell proliferation regardless of estrogen receptor status or estrogen sensitivity, and that its cellular targets include microtubules. In the present study, we investigated the apoptosis-inducing effects of A(II). MCF-7, MCF-7/LY2, and MDA-MB-231 cells all showed nuclear fragmentation in response to 100 microM A(II) when stained with Hoechst 33342 and examined by fluorescence microscopy. Pulsed field gel electrophoretic analysis showed that each of the cell lines also developed specific high molecular weight DNA fragments: a low level of 1-2 Mb fragments appeared after 6 hr, while 30-50 kb fragments accumulated subsequently. At 24 hr of drug exposure, the majority of cells became nonadherent, and the 30-50 kb fragments were restricted to detached MCF-7 and MDA-MB-231 cells. Both adherent and detached MCF-7/LY2 cells exhibited these fragments. A previous study by single-color (propidium) flow cytometry demonstrated that A(II) blocks MDA-MB-231 cells in G2/M of the cell cycle. More refined analyses in the present study showed this same result for MDA-MB-231 cells, but MCF-7 and MCF-7/LY2 cells did not reveal apparent drug-induced cell cycle block. A(II) demonstrated growth inhibitory, cell cycle-perturbing, and hypodiploidy-inducing activity against other human breast carcinoma lines, i.e. BT-20, CAMA-1, and SKBR-3, but no such actions in the non-tumorigenic, "normal" human breast epithelial line MCF-10A. Bromodeoxyuridine labeling and two-color flow cytometric analysis, however, suggested that A(II) caused stimulation into S phase, and that G2/M was the phase of the cell cycle from which cells apoptosed. A(II) caused cell rounding, detachment from the growth matrix, and nuclear shrinkage and fragmentation in parallel with biochemical changes. Cycloheximide inhibited A(II)-induced cell death, indicating that its toxicity requires de novo protein synthesis.     

High avidity CTLs for two self-antigens demonstrate superior in vitro and in vivo antitumor efficacy

A majority of the human tumor-associated Ags characterized to date are derived from nonmutated "self"-proteins. Little is currently understood about the nature of the self-reactive lymphocytes that recognize these Ags. We recently characterized two nonmutated tumor-associated Ags for the B16 murine melanoma: tyrosinase-related protein-2 (TRP-2) and the endogenous retroviral envelope protein, p15E. We previously reported that both TRP-2 and p15E reactive CTL could be detected in the spleens of naive animals after a single in vitro stimulation using 10(-5)-10(-6) M of the appropriate Kb-binding 9-amino acid epitope. In this report we show that the CTL found in naive animals are low avidity lymphocytes, that respond only to high concentrations of peptide in vitro. We demonstrate that titration of in vitro-stimulating peptide to limiting concentrations distinguishes qualitative differences in the lymphocyte reactivity to these two Ags between vaccinated and unvaccinated animals. We further demonstrate that in vitro expansion of CTL in either high or low concentrations of stimulating peptide generated CTL cultures with different avidities for the relevant epitopes. CTL expanded in low concentrations demonstrated higher avidity for peptide-pulsed targets and better tumor recognition, when compared to CTL generated in the presence of high concentrations of Ag. More importantly, high avidity CTL demonstrated superior in vivo antitumor activity. These results demonstrate that qualitative differences in the CTL that recognize these two self-Ags are critically important to their in vitro and in vivo anti-tumor efficacy.