Thrombosis and ELISA optical density values in hospitalized patients with heparin-induced thrombocytopenia.

The natural history of heparin-induced thrombocytopenia (HIT) in the absence of thrombosis was previously established using functional assays for confirmation of diagnosis (e.g. 14C serotonin release assay). An enzyme-linked immunosorbent assay (ELISA) that detects the presence of antibodies directed against the heparin-platelet factor-4 (PF4) complex has largely replaced functional assays in many medical centers. Although the ELISA is highly sensitive for detecting HIT antibodies, its usefulness for predicting thrombotic outcomes has not been clearly established. We performed a retrospective chart review of all hospitalized patients at a university hospital who tested seropositive for HIT by a commercial ELISA during 2001 and 2002. A total of 63 inpatients were identified as HIT positive by ELISA. Forty-eight patients had no apparent HIT-associated thrombosis at the time of HIT seropositivity (i.e. isolated HIT) and only one was treated prophylactically with a direct thrombin inhibitor. The 30-day thrombosis rate for patients with isolated HIT was 17% (eight of 48). Higher ELISA optical density (OD) measurements correlated significantly with thrombosis (1.41 +/- 0.87 vs. 0.79 +/- 0.46, P <0.001). Patients with isolated HIT and an OD measurement of > or = 1.0 demonstrated nearly a 6-fold increased risk of thrombosis compared with those with OD values between 0.4 and 0.99 (odds ratio 5.74, 95% confidence interval 1.73, 19.0; absolute rate of thrombosis, 36% vs. 9%, respectively, P=0.07). We conclude that in hospitalized patients with isolated HIT, the presence of heparin-PF4 antibodies detected by ELISA was associated with a significant risk of subsequent thrombosis and higher ELISA values were observed among patients suffering thrombotic events.

Possible association between the Norplant contraceptive system and thrombotic thrombocytopenic purpura.

BACKGROUND: Thrombotic thrombocytopenic purpura (TTP) is a rare, potentially fatal disease of uncertain etiology. Early diagnosis and treatment are essential to patient survival. The ]purpose of this report is to describe three patients with levonorgestrel implants (Norplant system) who developed TTP. CASES: A 24-year-old woman with levonorgestrel implants in place for 7 months was admitted to our hospital for treatment of TTP. Clinical symptoms included easy bruising, menorrhagia, headaches, and fever; laboratory evaluation revealed thrombocytopenia (18 x 10(9)/L) and microangiopathic hemolytic anemia. She was treated with plasmapheresis, and the implants were removed. Through the Freedom of Information Act, we reviewed all adverse events associated with Norplant use reported to the Food and Drug Administration (FDA) as of the end of 1992. Two additional cases were identified. CONCLUSIONS: Although a causal relationship between progestogen-only contraceptives and TTP is not established by the data presented, these three cases may represent an increased incidence of TTP in women using levonorgestrel implants. Patients who receive Norplant should be advised to seek medical attention if symptoms appear. Physicians and other health care providers should be aware of the possible association between use of the Norplant system and TTP and are urged to report similar cases to the FDA.

Transglutaminase activity and embryonal carcinoma cell differentiation.

Murine embryonal carcinoma (EC) cells induced to differentiate by retinoic acid (RA) modulate transglutaminase (TGase) activity shortly after exposure to the inducer. Compounds that inhibit TGase enzyme activity in vitro can successfully block RA induced EC cell differentiation in culture. These observations suggest that TGase may play a role in mediating RA induced EC cell differentiation.

alpha-Difluoromethylornithine induces differentiation of a human embryonal carcinoma cell line in vitro.

Human embryonal carcinoma cells could serve as a useful model system for analysis of early human development. A limited number of human embryonal carcinoma cell lines have been generated from in vivo tumors. We report here that alpha-difluoromethylornithine, a specific enzyme-activated inhibitor of ornithine decarboxylase activity, can induce differentiation in human embryonal carcinoma cells. The differentiated phenotype could be distinguished from undifferentiated cells by altered cellular morphology, biochemical and cell surface antigenic properties. These results suggest that alterations in the intracellular levels of polyamines may play a role in human embryonal carcinoma cell differentiation, and possibly human embryogenesis.

A comparative study of blood warmer performance.

Massive transfusions of refrigerator-temperature blood may induce hypothermia and life-threatening arrhythmias; for this reason a variety of devices have been developed for rapid blood warming. Blood warmers available in the United States use one of three warming technologies: dry heat, water bath, or countercurrent heat exchange. In the current study we evaluated blood warmers representative of each technology for speed and extent of heat transfer: the Fenwal blood warmer (Fenwal Laboratories; dry heat), the DW-1000 (American Pharmaseal Co.; dry heat), the FloTem IIe (DataChem Inc.; dry heat), the Hemokinetitherm (Dupaco Inc.; water bath), and the H250 and H500 (Level 1 Technologies; countercurrent heat exchange). Only one countercurrent heat instrument (the H500) was able to heat blood > or = 33 degrees C at target flow rates > or = 250 ml/min. Dry heat and water bath blood warmers were unable to warm blood > or = 33 degrees C at target flow rates > or = 100 ml/min. High resistance to flow with the proprietary tubing required for one instrument (the Hemokinetitherm) prevented tests of blood warming at rates > 150 ml/min. We found that instruments that used countercurrent technology warmed blood and saline more effectively than did blood warmers that used either dry heat or water bath technology. Our study also demonstrated the need for close control and standardization of experimental conditions in the evaluation of blood warming devices.

Unexpected citrate toxicity and severe hypocalcemia during apheresis.

BACKGROUND: The citrate anticoagulant used during apheresis procedures is considered a safe medication because it is rapidly metabolized by the donor. However, acute, life-threatening hypocalcemia is possible if the infusion rate of citrate is increased. CASE REPORT: A 54-year-old woman with metastatic breast cancer, but otherwise in good health, had just begun a fifth collection of hematopoietic peripheral blood progenitor cells by leukapheresis. The instrument's self-loading apheresis kit was primed uneventfully. Seven minutes into the procedure, the patient developed signs and symptoms suggesting severe hypocalcemia, including muscle spasms, chest pain, and hypotension. The citrate bag was discovered to have emptied, and a section of the anticoagulant tubing was protruding outside of the rotary pump. The patient's ionized calcium level was 0.64 mmol per L (normal range, 1.18-1.38 mmol/L). In subsequent experiments where the anticoagulant tubing was either improperly loaded at the outset or partially pulled out of the rotary pump, no instrument alarms sounded. CONCLUSION: Citrate toxicity and life-threatening hypocalcemia can occur if the anticoagulant line of an apheresis instrument is not properly housed in its rotary pump or becomes disengaged during the procedure. Instrument manufacturers are encouraged to consider designs that allow the direct measurement of the volume of citrate delivered. In the interim, periodic visual and tactile confirmation of tubing placement during apheresis procedures is prudent.

A multicompartmental model of vitamin A kinetics in rats with marginal liver vitamin A stores.

A linear, first-order, constant-coefficient multicompartmental model is presented which describes the dynamics of [3H]retinol turnover in adult rats with normal plasma retinol concentrations but low liver stores (less than 100 micrograms of retinol equivalents). To fit plasma and tissue (liver, kidney, and rest of carcass) tracer and tracee data, eight physiological compartments were required in the model: two in plasma (proposed to correspond to the retinol transport complex, and retinyl esters in plasma lipoproteins) and two each in liver, kidneys, and other extrahepatic tissues. Extensive recycling of retinol among plasma, liver, and the rest of carcass was also required. The model predicted that 44% of whole body vitamin A (143 micrograms) was in extrahepatic tissues. The vitamin A utilization rate (system disposal rate) was 6.9 micrograms of retinol equivalents/day. The system residence time (mean sojourn time) for vitamin A was 21 days, and the fractional catabolic rate for the system was 5%/day. The mean transit time (turnover time) for vitamin A in its plasma retinol transport complex was 0.078 days (1.9 hr); the residence time was 0.98 day, versus 11 days in the liver, 9 days in carcass, and 0.54 days in kidneys. The model predicted that, of the plasma turnover, 48% recycled to the liver and 52% to extrahepatic tissues. The liver retinol secretion rate was 48 micrograms/day, more than half of which was from recycled plasma retinol. Since the plasma retinol turnover rate (87 micrograms/day) was 13 times the system disposal rate, the data suggest that this is a high response system in which changes in the dynamics of recycling of retinol allow for rapid adjustment in vitamin A distribution in response to changes in nutritional, metabolic, or physiological conditions; and in which plasma retinol levels are controlled homeokinetically by changes in hepatic and extrahepatic recycling of holo retinol-binding protein.

Hemolytic disease of the newborn due to the Scianna antibody, anti-Sc2.

BACKGROUND: Alloantibodies to the low-frequency antigen Scianna 2 (Sc2) are uncommon and not previously reported to cause immune hemolysis. CASE REPORT: A group B, Rh-negative infant born to a group B, Rh-positive mother had a 2+ direct antiglobulin test, as well as modest hyperbilirubinemia and a hematocrit of 45 percent. Ongoing immune hemolysis led to a hematocrit of 17.3 percent on Day 20 of life, and the infant required hospitalization and red cell transfusions. The routine maternal antibody screen was negative, but anti-Sc2 was detected during work-up for a low-frequency red cell antigen, and the father's red cells typed as Sc:1,2. CONCLUSION: Anti-Sc2 can cause clinically significant hemolytic disease of the newborn. Although the antibody is uncommon, its frequency and hemolytic potential may be underappreciated, in part because investigations often are not carried out in the infant whose red cells are ABO-incompatible with maternal blood.

Mycobacteremia and granulomatous hepatitis following initial intravesical bacillus Calmette-Guerin instillation for bladder carcinoma.

We describe the first case of mycobacteremia and granulomatous hepatitis occurring after the initial intravesical instillation of bacillus Calmette-Guerin (BCG) for bladder cancer. Eight days after BCG instillation, a liver biopsy revealed well-defined granulomas and acid-fast bacilli. A blood culture drawn 8 h after BCG instillation grew Mycobacterium bovis. We summarize the reported complications of BCG intravesical immunotherapy, the associated risk factors, and discuss options for treatment. Although rare, mycobacteremia and granulomatous hepatitis are important systemic side effects of intravesical instillation of BCG for bladder cancer, and should be considered in any patient who presents with persistent fever and abnormal liver function tests after instillation of BCG.

Lowering the hemoglobin cutoff for female plateletpheresis donors.

BACKGROUND: The standards of the American Association of Blood Banks describe a minimum hemoglobin level of 12.5 g per dL for apheresis donors. Until 1995, the authors' institution accepted occasional platelet donors with a lower minimum hemoglobin (11.5 g/dL), if accompanied by medical director approval. STUDY DESIGN: All donation records from a 6-month period before 1995 were retrospectively reviewed to determine whether this lower hemoglobin cutoff adversely affected either the safety of the platelet donation process or donors' subsequent hemoglobin levels. RESULTS: Of 450 donations, 56 (12%, Group 1) were from donors with hemoglobin concentrations between 11.5 and 12.4 g per dL (2 donations from 1 man; 54 donations from 45 women). The remaining 394 donations (88%, Group 2) came from donors with hemoglobin concentrations > or = 12.5 g per dL (216 donations from 118 men; 178 donations from 119 women). The frequency of donor reactions was acceptable (Group 1, 11%; Group 2, 6%); 2 percent of donations by Group 1 donors and 1 percent by Group 2 donors were terminated because of these reactions. Of 46 donors in Group 1, 30 returned to donate platelets again at a later time; at least once, 23 (77%) had a hemoglobin > or = 12.5 g per dL. Ten donors in Group 1 returned for additional donations within 56 days; no meaningful decrease in hemoglobin levels occurred. A hemoglobin cutoff of 12.5 g per dL during the study period would have excluded 1 percent of platelet donations by men and 23 percent by women. CONCLUSION: The data demonstrate that the lower hemoglobin cutoff of 11.5 g per dL is a safe and relevant threshold for accepting female plateletpheresis donors and would allow more participation by women in blood donor programs.

The collection and evaluation of peripheral blood progenitor cells sufficient for repetitive cycles of high-dose chemotherapy support.

BACKGROUND: The development of an optimized peripheral blood progenitor cell (PBPC) harvest protocol to provide support for repetitive chemotherapy cycles is described. STUDY DESIGN AND METHODS: PBPCs mobilized by cyclophosphamide plus granulocyte-colony-stimulating factor (G-CSF) were studied in 163 leukapheresis harvests from 26 lymphoma patients. Harvested cells were transfused with two chemotherapy cycles and with an autologous bone marrow transplant. Progenitor cell content was examined in the context of hematopoietic engraftment. RESULTS: Mobilization allowed the harvest of large numbers of PBPCs. Peak harvests tended to occur after the recovering white cell count exceeded 10 x 10(9) per L. CD34+ lymphomononuclear cell (MNC) and colony-forming units-granulocyte-macrophage (CFU-GM) counts correlated poorly, but both measures peaked within 24 hours of each other in 21 of 26 patients, which demonstrated PBPC mobilization. Engraftment of platelets (> 50 x 10(9)/L) and granulocytes (> 500 x 10(6)/L) was achieved in a median of 20.5 and 16 days, respectively. A minimum number of progenitors necessary to ensure engraftment could be derived. CONCLUSION: Cyclophosphamide and G-CSF allowed the harvest of sufficient PBPCs to support multiple rounds of chemotherapy. Harvest should commence when the recovery white cell count exceeds 10 x 10(9) per L. PBPC harvest CD34+MNC counts are as useful as CFU-GM results in the assessment of PBPC content, and they may allow harvest protocols to be tailored to individual patients.

Liquid nitrogen freezers: a potential source of microbial contamination of hematopoietic stem cell components.

BACKGROUND: The recent report of hepatitis B transmission between hematopoietic progenitor and putative stem cell (HPC) components stored in liquid nitrogen led to the questioning of whether evidence existed for similar contamination by bacterial or fungal elements. STUDY DESIGN AND METHODS: Microbial contamination rates were reviewed for 704 HPC components from 255 patients over an 18-month period. Five liquid nitrogen freezers were surveyed for microbial contamination. The literature was reviewed to ascertain the published experience of other laboratories with HPC component contamination first documented on thawing. RESULTS: Seven (1.2%) of 583 thawed components were found to be contaminated with a variety of environmental or waterborne organisms, despite a meticulous protocol to prevent contamination during thawing. All of these components had been sterile on cryopreservation. Literature review revealed a similar incidence of post-thaw contamination from other centers. Microbial survey of liquid nitrogen freezers revealed low-level contamination in four of five. The organisms represented were similar to those cultured from thawed HPC components. One freezer was heavily contaminated by Aspergillus species. CONCLUSION: Liquid nitrogen freezers are not sterile, and both the liquid and vapor phases are potential sources of microbial contamination of HPC components. While low-level contamination by environmental organisms may be common, the occurrence of heavy contamination by potential pathogens such as Aspergillus species suggests that monitoring of liquid nitrogen sterility may be indicated. Strategies to assess and prevent microbial transmission from liquid nitrogen to HPC components need further development.

Preapheresis peripheral blood CD34+ mononuclear cell counts as predictors of progenitor cell yield.

BACKGROUND: Peripheral blood progenitor cells, harvested by apheresis after mobilization, provide rapid hematologic recovery after high-dose chemotherapy. However, because harvesting these cells is expensive and time-consuming, there has been much interest in optimizing collection protocols. An investigation was made to determine whether, in this clinical setting, peripheral blood progenitor cell yields may be predicted from preapheresis progenitor cell counts, allowing the length of each procedure to be "fine tuned" to achieve specific target goals. STUDY DESIGN AND METHODS: Preapheresis peripheral blood CD34+ cell and total colony-forming cell counts were assessed before 78 peripheral blood progenitor cell collections from 13 consecutive patients were performed. Preapheresis counts were correlated with actual progenitor cell yields. Factors affecting this correlation were analyzed. RESULTS: With the use of linear regression analysis preapheresis progenitor cell counts were found to correlate significantly but weakly with actual yields per kg of body weight per liter of blood processed (CD34+ cells: r = 0.43; colony-forming cells: r = 0.56). Further analysis revealed two possible causes: 1) circulating progenitor cell concentrations fluctuate widely during harvest, which implies that preapheresis counts are not representative of actual concentrations during apheresis, and 2) the efficiency with which apheresis machines extract mononuclear cells varies greatly between procedures. CONCLUSION: Preapheresis CD34+ and colony-forming cell counts correlated poorly with subsequent yields in this clinical setting, which suggests that it is not practical to use such counts to predict with certainty the length of apheresis needed to achieve a target yield.

Distribution of oral Haemophilus species in dental plaque from a large adult population.

The periodontal status of maxillary first molars in 284 young adults demonstrating near-health to early disease was evaluated, and supragingival and subgingival plaque samples were collected. Plaque samples were processed anaerobically, enumerated microscopically for bacterial morphotypes, and cultivated on various media to enumerate the microflora. Although haemophili were ubiquitous (recovered in 98.5 and 96.2% of the supragingival and subgingival plaque samples, respectively), 50% of the respective samples had proportions of less than or equal to 1.5% and less than or equal to 0.33% total Haemophilus spp. based on total cultivable microflora. To study the distribution of Haemophilus spp., 377 colonies were identified from modified chocolate agar (selective for oral haemophili) from 14 supragingival and corresponding subgingival samples from 14 subjects. The most prevalent species, Haemophilus parainfluenzae, was found in significantly higher proportions, based on total haemophili on modified chocolate agar, in supragingival and subgingival samples from teeth with shallower probing depths (less than or equal to 3.0 mm) versus deeper probing depths (greater than or equal to 3.0 mm). Additional statistically significant findings included Haemophilus segnis in higher proportions in supragingival samples from deeper sites, Haemophilus aphrophilus in higher proportions in subgingival samples from deeper sites, and Haemophilus paraphrophilus in higher proportions in subgingival samples from shallower sites. Scatter diagrams illustrating the bivariate distributions of proportions of haemophili with proportions of dark-pigmented Bacteroides spp., spirochetes, and streptococci demonstrated that high proportions of haemophili were never recovered from sites with high proportions of Bacteroides spp. or spirochetes. All levels of haemophili, however, were recovered from sites with all levels of streptococci. Two potential systems for interpreting haemophili data were hypothesized for predicting periodontal probing depths. There was highly significant agreement between the two systems. Small but statistically significant correlations were found between the gingival index, probing depth, and attachment level, and proportions of total Haemophilus species in the respective samples.

Rituximab and ifosfamide, mitoxantrone, etoposide (RIME) with Neupogen support for B-cell non-Hodgkin's lymphoma prior to high-dose chemotherapy with autologous haematopoietic transplant.

A phase I/II study was performed to analyse the ability of ifosfamide-based chemotherapy with rituximab to produce a turmour-free graft as well as the safety of retuximab prior to stem cell harvest and post high-dose chemotherapy. Twenty-two patients with B-cell non-Hodgkin's lymphoma were enrolled either having aggressive large-cell disease in relapse or at high/high-intermediate risk of relapse, or refractory lymphoma or mantle cell lymphoma, or indolent lymphoma. Chemotherapy consisted of ifosfamide 2 g/m2, days 1-3 with mesna, etoposide 100 mg/m2, days 1-3, and mitoxantrone 8 mg/m2 day 1, with figrastim. Rituximab was given at 375 mg/m2 for 4 doses. An encouraging overall response rate of 90%, including 11 CRs was achieved. CD34+ cells were successfully mobilized in 18 or 19 patients analysed so far with a median number of 3.4 x 10(6) cells/kg. The combination of ifosfamide-based chemotherapy with rituximab significantly reduced the number of contaminating B-cells in the stem cell product and so far there has only been a single relapse post high-dose chemotherapy with autologous haematopoietic transplant. The RIME regimen was generally well tolerated with minimal non-haematological toxicity and most of the treatment was done completely on an outpatient basis. Haematological toxicity was manageable with filgrastim, there were some infectious complications.

A phase I-II study of rituximab, ifosfamide, mitoxantrone and etoposide (R-IME) for B cell non-Hodgkin's lymphoma prior to and after high-dose chemotherapy and autologous stem cell transplantation (HDC-ASCT).

This phase I-II study describes the safety of rituximab, ifosfamide, mitoxantrone and etoposide (R-IME) as an induction regimen prior to high-dose chemotherapy and autologous stem cell transplantation (HDC-ASCT), and rituximab given post-HDC-ASCT for B cell non-Hodgkins's lymphoma. This study also measured the effect on disease burden and stem cell contamination. Patients with relapsed, refractory or poor risk B cell lymphomas were eligible. Patients were treated with two cycles of R-IME; all non-progressing patients under-went a third cycle and peripheral blood stem cell (PBSC) collection. Patients underwent HDC-ASCT and those patients in remission after HDC-ASCT were treated with four additional doses of rituximab. Tumor cell contamination was measured at baseline and in the PBSC. Serial immunoglobulin levels were measured. Patients were followed for time to treatment failure (TTF) and overall survival (OS). Thirty-two patients were enrolled. Thirty patients had at least stable disease after two cycles of R-IME. Twenty-nine underwent stem cell collection. The response rate to R-IME induction was 77% (20/26) with 35% (9/26) complete response(CR). Stem cell mobilization was successful in 93% (27/29) of patients. The response rate to R-IME induction and HDC-ASCT was 95% with a confirmed CR of 68%. Median follow-up was 28 months; the median TTFand OS have not been reached. There was a significant decline in stem cell tumor cell contamination and a significant decline in IgG without an increase in infections. Forty-three per cent of patients had transient neutropenia after post-transplant rituximab. R-IME is an effective cytoreductive and mobilization regimen. There appears to be a reduction in the number of lymphoma cells in the stem cell product and the toxicity is manageable.