Variant forms of procoagulant-like factor VIII in hemophiliacs.
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Animals↗
Biomedical subjects
G Rock
Publications and source records attributed to G Rock.
Changes in the Canadian blood system: the Krever Inquiry, Canadian Blood Services and Héma-Québec. Krever Commission.
Beginning in 1992, Canada underwent a series of investigations into the national blood program. The Krever Commission issued an interim report in 1993 and a final report in 1997. In all a total of 93 recommendations were made. Subsequently, in 1998, the Canadian Red Cross Society (CRC) withdrew from the blood program and two new organizations, the Canadian Blood Services (CBS) and Héma-Québec (HQ) took over as operators.
Blood Banks↗
Metabolic changes during platelet storage.
Platelet concentrates (PC) were stored in plastic bags with continuous shaking at 4, 22, and 37 C. Various metabolic parameters were examined over a 72-hour period. At 22 C, the pH and PO2 declined over 72 hours while the PCO2 and lactate increased. Hypotonic shock declined to 70 per cent. This differed from the small amounts of CO2 and lactate found at 4 C and the marked accumulation of metabolites and almost complete loss of shock response at 37 C. Aggregation was always better maintained with 4 C storage. The toxic effect of the accumulation of metabolites on the platelets was tested by adding lactate to fresh PC at zero time. This was effective in lowering the initial pH, markedly inhibiting the response to aggregation and decreasing the total accumulation of lactate during storage, but did not produce an inhibition of hypotonic shock response. The effect of accumulation of toxic metabolites was further investigated by using 72-hour plasma and platelets and recombining each of them with fresh preparations. Platelets were tested under degassed conditions to outline the requirements for oxygen and gasious exchange. Surprisingly, there was less accumulation of lactate and CO2 and better hypotonic shock response. These experiments have detected various changes in viability markers in platelets that are stored under actual blood bank conditions and indicate that the accumulation of lactate is not totally responsible for the toxic inhibition of platelet performance that is found upon storage at 22 C.
Blood Cell Count↗
Variations in cryoprecipitate production.
Several variables influencing the recovery of Factor VIII in the cold-precipitated protein fraction were examined. The actual rate at which the physical-chemical conversion of plasma from a solid to a liquid state occurs does not seem to affect the yield. Rather, it is the amount of time immediately postthaw and prior to centrifugation that determines the Factor VIII activity in the cryoprecipitate. With prolonged periods at 4 C the Factor VIII activity leaves the cold precipitate and lifts into the supernatant. Even distribution of the plasma layer and close attention to the thawing procedure facilitate Factor VIII recovery.
Chemical Precipitation↗
LISS--an effective way to increase blood utilization.
The low ionic strength solution (LISS) of Low and Messeter was compared with both the automated low ionic strength Polybrene and the manual IDAT techniques. A five minute incubation with the LISS was sufficient to detect all significant antibodies. By extending the incubation period to 15 minutes it was possible to increase the sensitivity of the reaction (as measured by titer) beyond that of either of the other methods. This LISS procedure has enabled us to greatly extend the applications of a "standby procedure" for elective surgery. In this procedure routine crossmatching is not done. Rather, the blood is placed on standby and if required, transfusion is provided by using the LISS. In one general hospital this resulted in the reduction by 1,600 units of unneccessary crossmatches and an increase of 10% in the total blood utilization rate over a nine-month period.
Antibodies↗
The effect of aspirin on the hypotonic shock response.
The effect of aspirin on the hypotonic shock response was studied at 2, 6, 24, and 48 hours after ingestion of 1.6 g of acetylsalicylic acid. There was a marked acceleration of the recovery phase of the response at two hours. This steadily decreased and by 48 hours had returned to near normal values. In most cases there was also a change in the initial phase of the shock response. We have interpreted these results to indicate a change in membrane permeability (first phase effect) as well as an activation of the energy-dependent recovery (second phase). The latter process possibly is the result of an increase in the levels of available ATP. As previously reported, the response to aggregating agents was impaired in the presence of ASA. Normal and aspirinated platelets were mixed in an attempt to overcome the ASA effect. Using two hour postingestion samples we noted considerable impairment of both the hypotonic shock response and aggregation even in the presence of very high concentrations of normal platelets.
Aspirin↗
The accumulation of mono-2-ethylhexylphthalate (MEHP) during storage of whole blood and plasma.
The accumulation of the plasticizer di-2-ethylhexylphthalate (DEHP) in blood and blood components has been of considerable concern for some time. We have followed the accumulation of DEHP and one of its major metabolities, mono-2-ethylhexylphthalate (MEHP) during storage of whole blood, platelet-rich plasma, platelet concentrates, and platelet-poor plasma for periods ranging from 72 hours to four weeks. Both phthalates showed a progressive increase in concentration with time. While the levels of DEHP were much greater than those of MEHP, there was nonetheless a significant and continual increase in MEHP in all preparations. The highest concentrations of both DEHP and MEHP were found in the platelet-poor plasma, indicating that platelets do not have a major role in the accumulation of the phthalates in blood. The accumulation of MEHP was shown to be a direct result of the metabolism of DEHP by plasma protein(s) rather than leaching from the blood bag.
Blood Platelets↗
Platelet storage: an assessment of the requirements for plasma and oxygen.
Platelets from several units of plasma were pooled, and then resuspended either as platelet concentrate (PC) (60 ml) or as platelet rich plasma (PRP) (220 ml) and followed during 72 hours of storage at 22 C. Aggregation, pH, and hypotonic shock response are better maintained in the larger volume of plasma. However the decreased pH and function in the PC is not the result of lactate production. While the relative concentration, expressed in mg/dl, is higher in the PC, the absolute production per platelet and, therefore, the total amount of lactate is higher in PRP. Glucose levels are always sufficiently high to permit glucose to be used as a metabolic substrate even after 72 hours. PC maintained in nitrogen have better in vitro function than do platelets stored either in oxygen or CO2. CO2 can easily diffuse through the plastic bag; the same is not true of oxygen. Less than 15 per cent of the volume of oxygen introduced into the bag passed out through the PVC plastic over a 72-hour period. The data suggest that during storage, anaerobic glycolysis is the preferred metabolic route. The relatively poor performance of the PC maybe the result of limitation of some unidentified substrate or cofactor which is present in plasma and which is necessary for maintaining function.
Blood Platelets↗
A new technique for the collection of plasma: machine plasmapheresis.
The Haemonetics Model 50 permits the collection of 500 ml of plasma within an average of 30 minutes, and the donor is never disconnected from his cells. In a detailed assessment of 28 donors, we found no detrimental effect of the procedure. There was no evidence of fibrin split products or complement activation. The plasma showed good recovery of protein with slightly elevated factor VIII levels; citrate levels are only two-thirds of manual plasmapheresis values. While there was a relatively large number of platelets collected into this plasma, the platelets were small, with a mean diameter of 1.8 mu and poor response to aggregation. Therefore it would appear that this plasma should not be used to make a platelet preparation. Nonetheless, evaluation of this machine indicated that the performance parameters are acceptable and that donor acceptance is exceptional, with widespread enthusiasm for this "new" method of blood donation.
Blood Donors↗
Plasma exchange in the treatment of hemolytic disease of the newborn.
Plasma exchange has become a widely used therapeutic tool in the treatment of immune-mediated disease. This technique has been applied to the treatment of gravid women whose high antibody titers and past history of stillbirths indicate a significant possibility of fetal loss due to HDN. The procedure was effective in decreasing both antibody titer and the quantity of antibody in four of five patients. In one other case, the antibody titer rose toward the end of the pregnancy; however, the quantity of anti-D remained low. The IgG:IgM ratio and the subclass specificity of IgG did not significantly change with treatment. Hematologic monitoring also indicated no adverse effects, and three of the five women delivered viable fetuses. Two of the other women produced infants only moderately affected by HDN but stillborn within days of intrauterine transfusion. The results indicate that plasma exchange is a useful therapeutic consideration in the sensitized gravid women.
Amniocentesis↗
The effect of adenine on platelet storage.
Adenine is an approved additive to citrate-phosphate-dextrose anticoagulant for whole blood collection and extends the storage life of red blood cells. We used in vitro methods to investigate the effects of adenine on platelet function and viability during 72 hours of storage at 22 degrees C. Although the hypotonic shock response and aggregation were decreased, these effects were reversed following separation and resuspension in fresh adenine-free plasma. Serotonin uptake and release were not affected by adenine, however malonaldehyde formation was slightly enhanced. Glucose, pH and pO2 levels were lower, while lactate levels were slightly higher than in platelets stored without adenine. These results indicate little in vitro effect of adenine on platelets stored as platelet-rich plasma or platelet concentrate.
Adenine↗
Automated platelet production during plasmapheresis.
Manual plasmapheresis is widely used to permit collection of fresh-frozen plasma with return of the red cells to the donor. However, this method is time-consuming and carries the inherent risk of returning the wrong cells to any individual donor. Automated plasmapheresis, using a specially designed discontinuous cell separator, permits the collection of 500 ml of plasma within 30 minutes without disconnecting the donor from the bag containing the donor's red cells. However, this plasma contains a small number of platelets which are not suitable for transfusion. We now report the modification of this machine to permit simultaneous collection of 3 units of platelet concentrate as well as 500 ml of plasma in less than 50 minutes. Reduction of the g force of the separator from 1200 to 650 g permits the simultaneous collection of plasma and a platelet concentrate with an average yield of 2.21 X 10(11) platelets. Platelet size distribution, in-vitro function. 51Cr survival, posttransfusion increment, and bleeding time correction are all normal. This modification has increased the flexibility of the separator and provides an instrument which can be used with a random volunteer blood donor population to generate both plasma and cells in less than 50 minutes and with increased cost-effectiveness over plasmapheresis alone.
Automation↗
Rapid controlled thawing of fresh-frozen plasma in a modified microwave oven.
A microwave oven has been specifically modified to permit rapid thawing of fresh-frozen plasma (FFP) by using a rotating disc with a temperature sensor to hold the plasma bag. This modification makes it possible to mix the FFP continuously during thawing, and automatically shuts the oven off when the plasma reaches 21 degrees C. Comparisons were made between FFP thawed in the modified microwave oven and FFP thawed conventionally in a 37 degrees C waterbath. The following tests were done: total protein, albumin, and immunoglobulin concentrations; plasma fibrinogen, factor VIII, and factor IX activities; protein electrophoresis, albumin aggregation, hemolytic complement activity, and plasma particle count and size. In no case was there a significant difference between plasma thawed in the microwave oven compared with that thawed in the waterbath. Further, microwave thawing was reliable and rapid; all units of FFP thawed in less than 6 minutes, and the thawed plasma did not vary by more than 6 degrees C from the preselected final temperature of 21 degrees C. Thus, it appears that controlled thawing of FFP in a microwave oven specifically designed for this purpose is an effective and reliable method and has many advantages over conventional thawing of FFP.
Adenine↗
Modified fluid gelatin in leukapheresis accumulation and persistence in the body.
Current methods for centrifuged granulocyte procurement involve the use of an agent to produce red cell rouleaux and enhance separation of leukocytes. Hydroxyethyl starch (HES), the agent most frequently used, has the disadvantage of causing progressive volume expansion and persisting in the circulation for long periods. We therefore assessed modified fluid gelatin (MFG) as a possible replacement for HES during granulocyte collection. We found that MFG is cleared more rapidly from the circulation with no traces remaining 7 days after multiple exposure, as determined by hydroxyproline measurement. However, after four consecutive daily infusions, we measured 0.60 liters plasma expansion in four individuals tested, somewhat lower than the 0.85 liters previously reported for HES. Modified fluid gelatin causes no impairment of coagulation with normal prothrombin time (PT), partial thromboplastin time (PTT), and platelet function.
Cell Separation↗
Five-day storage of platelet concentrates.
The short 72-hour shelf-life of platelet concentrates stored in standard PL146 (Fenwal) plastic bags often results in shortages of platelets. This 3-day limitation is based on the biochemical and physiological changes that occur during storage and that result in decreased viability and survival after transfusion. We assessed both in vitro and in vivo function of platelet concentrates stored for 3 and 5 days in two new plastic packs: PL732 (Fenwal) and CLX (Cutter). The concentrate pH was maintained above 7.0 in both bags and there was little change in platelet count or size following 5 days of storage. Aggregation response to adenosine diphosphate, epinephrine, and collagen was maintained well. The PCO2 values indicated good gas escape with lower values after 5 days of storage than at 0 time. Lactate accumulation and glucose utilization were also lower in these new bags. Autologous survivals of chromium-labeled platelets stored for 5 days were 6.0 days (PL732) and 5.1 days (CLX), which are equal to or better than those found for platelets stored for 3 days in PL146. Posttransfusion increments in thrombocytopenic patients were acceptable; 49 percent after 1 hour and 31 percent after 24 hours for concentrates stored in CLX and 44 percent after 1 hour and 28 percent after 24 hours for concentrates stored in PL732. Both of these new bags, which contain different types of plasticizers, provide an environment that results in an improved product and will permit 5-day storage of platelet concentrates; these two benefits will help to alleviate the difficulties in supply of platelet concentrates.
Blood Platelets↗
Incorporation of plasticizer into red cells during storage.
The development of flexible plastic blood bags has permitted effective blood component production and therapy. However, the plasticizer di(2-ethylhexyl)phthalate (DEHP), whose toxicity in humans is still undefined, is known to leach from the plastic into stored blood. Despite the availability of bags made of plastics not using DEHP, the collection and storage of red cells is still done in DEHP plasticized packs, and in fact the storage life for red cells has recently been increased up to 49 days using new anticoagulant-preservative solutions. We examined the relationship between DEHP and stored red cells. We found that 28 percent of available 14C-DEHP binds immediately to sites in both the membrane and cytosol fractions of the red cells, and that the total amount and distribution of 14C-DEHP does not change significantly over 7 days. When red cell concentrates were stored with or without DEHP, using either plastic (polyolefin) bags not containing DEHP or glass, definite reduction in the osmotic stability of the red cells was found in the absence of DEHP. Plasma-free hemoglobin levels were 90.3 mg per dl after 35 days of storage in plastic packs containing DEHP and 181.7 mg per dl in the polyolefin bags. The advantages of improved in vitro stability of red cells stored in plastics containing DEHP must be weighed against the potential hazards of patient exposure to DEHP.
Blood Preservation↗
Simple and rapid preparation of granulocytes for the treatment of neonatal septicemia.
Recent reports have documented benefit from the infusion of granulocytes into septic neonatal patients. However, while the requirement of 1 X 10(9) per kg of granulocytes is easily met by apheresis procedures, treatment of neonatal septicemia requires immediate response 24 hours a day. Unfortunately, this rapid response is beyond the capability of most apheresis units. Therefore, we developed a method for obtaining granulocytes from whole blood collected by routine blood donation. Hydroxyethyl starch (60 ml of a 6% solution) is introduced into a unit of whole blood within 24 hours of collection. Following sedimentation and subsequent centrifugation, a buffy coat is obtained which contains more than 75 percent of the granulocytes; an average of 1.62 X 10(9) WBC, of which 1.25 X 10(9) were granulocytes that were recovered in an average 17-ml final volume with a hematocrit of 4 percent. Granulocyte morphology and function as measured by chemiluminescence indicated good maintenance of function. Total processing time was short, requiring less than 2 hours, and the procedure was performed easily in a routine blood bank. The recovery of 1.25 X 10(9) granulocytes from a single, routine blood donation without steroid treatment of the donor makes this product ideal for use in the treatment of neonatal septicemia.
Cell Separation↗
Factors that influence lymphocyte yields in lymphocytapheresis.
The recent use of lymphocytapheresis to treat immune-mediated diseases such as rheumatoid arthritis prompted a study of factors that influence the cell composition of lymphocytapheresis concentrates. Following single cytapheresis procedures, using protocols recommended by manufacturers, lymphocyte yields were significantly higher with the model 2997 (IBM) and CS-3000 (Fenwal) cell separators as compared to the model 30 (Haemonetics) separator (9.8 +/- 1.1 and 7.1 +/- 1.2 X 10(9) lymphocytes, respectively, versus 4.6 +/- 1.1 X 10(9); p less than 0.01). The lymphocyte concentrate obtained with the CS-3000 separator contained the smallest number of monocytes (0.6 +/- 0.4 X 10(9) versus 1.4 +/- 1.6 and 1 +/- 0.3 X 10(9) for the model 2997 and 30 cell separators, respectively). Platelet contamination of the lymphocyte concentrate was highest with the CS-3000 (6.5 +/- 2.4 X 10(11], and erythrocyte contamination was highest when the model 30 was used (21 +/- 3.0%). Studies using the model 2997 indicated that lymphocyte yields were significantly influenced by donor pre-apheresis absolute lymphocyte counts, and for this cell separator by specific operating variables, such as channel centrifugation speed and positioning of the red cell interface during lymphocyte collection. Maximal yields were obtained when the channel centrifugation speed was 800 to 1000 rpm (equivalent to 100-150 X g) and the red cell interface was adjusted to yield a cell concentrate with a hematocrit less than 4 percent. These results suggest that it will be necessary to standardize lymphocytapheresis collection protocols in future studies to assess the role of lymphocytapheresis in the management of immune-mediated diseases such as rheumatoid arthritis.
Adult↗