Comparison of platelet indices in dengue fever patients based on platelet transfusion: A prospective observational study in a tertiary care center.

CONTEXT: Dengue fever is the most prevalent mosquito-borne viral disease in humans. Platelet indices (PIs) are given by the cell counters but are often not reported which is possibly due to under-recognition of the utility of these parameters. AIMS: This study aimed to compare PIs in patients with dengue fever to assess their role in the outcome such as hospital stay and platelet transfusion requirements. SETTINGS AND DESIGN: Prospective observational study in a tertiary care center, Thrissur, Kerala. SUBJECTS AND METHODS: A group of 250 dengue patients was studied over a period of 18 months. The platelet parameters (platelet count, mean platelet volume [MPV], platelet distribution width [PDW], platelet large cell ratio [PLCR], plateletcrit [PCT] and immature platelet fraction [IPF]) were measured with Sysmex XN-1000 and followed up every 24 h. The clinical features, duration of hospital stay and platelet transfusion requirements details were collected. STATISTICAL ANALYSIS USED: Independent t-test, Chi-square test, Karl Pearson correlation coefficient. RESULTS: A total of sample size was 250. The study showed normal PDW and MPV, low platelet count and PCT, and high PLCR and IPF in dengue patients. There were significant differences in PIs (lower platelet count and PCT, higher MPV, PDW, PLCR, and IPF) in comparison between dengue patients based on platelet transfusion. CONCLUSIONS: PIs may act as a predictive tool in the diagnosis and predicting outcomes in dengue fever. Low platelet count and PCT, high PDW, MPV, PLCR, and IPF in transfused dengue patients were found to be statistically significant. Clinicians need to be sensitized about the utility and limitations of these indices and rationalize the need for red cell and platelet transfusions in dengue.

WBC content of platelet concentrates prepared by the buffy coat method using different processing procedures and storage solutions.

BACKGROUND: The number of WBCs in platelet concentrates (PCs) prepared by the buffy coat (BC) method with different storage solutions can result in low (5 x 10(6)/unit) WBC levels by the use of careful centrifugation techniques without filtration. At present, most blood banks use filtration steps to meet these requirements. The difference in processing methods and suspension solutions prompted the investigation of the influence of the various procedures on the WBC and platelet content of PCs. STUDY DESIGN AND METHODS: PCs from 5 BCs were harvested without or with inline filtration (AutoStop BC, Pall Corp.) in either plasma (PCs-plasma) or platelet additive solution (PCs-PAS-2). After preparation, samples were taken for counting WBCs and platelets and for analyzing WBC subsets by flow cytometry using specific MoAbs. The WBCs were concentrated before analysis of the WBC subsets. Results less than 2.5 cells per microL were considered below the limit of accuracy of the subset analysis. RESULTS: All filtered PCs met the AABB standard of 5 x 10(6) per unit and the European guidelines of 1 x 10(6) per unit. None of the nonfiltered PCs met the European guidelines, but all met the AABB guidelines. All filtered units gave residual WBC counts below the detection limit for subset analysis. Filtered PCs-plasma gave significantly higher platelet counts than filtered PCs-PAS-2 or nonfiltered PCs (p<0.01, ANOVA). CONCLUSION: Careful centrifugation of pooled BCs, with plasma or PAS-2, can result in PCs with low WBC contamination levels. However, filtered PCs are superior, because of better WBC removal and higher platelet counts.

Reactions and platelet increments after transfusion of platelet concentrates in plasma or an additive solution: a prospective, randomized study.

BACKGROUND: Reactions after platelet transfusions are rather common and frequently are caused by plasma constituents. In recent developments, the preparation and storage of platelet concentrates (PCs) in a platelet additive solution (PAS-2) have been shown to result in acceptable storage conditions. A major drawback of the use of these PCs is the progressive increase of P-selectin-positive platelets during storage. The clinical benefit of transfusions of PCs in PAS-2 was studied. STUDY DESIGN AND METHODS: PCs prepared from buffy coats were suspended in either plasma or PAS-2 and stored for up to 5 days. Clinical responses were evaluated in a prospective study in 21 patients treated with intensive chemotherapy for hematologic malignancies. Eligible patients were randomly assigned to receive prophylactic transfusions of PCs prepared in either plasma or PAS-2. Reactions and CCIs were recorded after each transfusion. RESULTS: The incidence of reactions in 12 patients given PCs in plasma (n = 192) was 12 percent. Transfusions to 9 patients of PCs in PAS-2 (n = 132) showed a reduction in the incidence of reactions to 5.3 percent (p<0.05). The average 1-hour and 20-hour CCIs after transfusion of PCs in plasma were 20.7 +/- 8. 5 and 11.5 +/- 8.0, respectively. CCIs after transfusion of PCs in PAS-2 were significantly lower: the average 1-hour CCI was 17.1 +/- 6.6 (p<0.001) and the average 20-hour CCI was 9.5 +/- 7.0 (p<0.05). Storage conditions of PCs were optimal: in each group, average 1-hour CCIs of both fresh and stored PCs were similar. The 20-hour CCIs after the transfusion of fresh and stored PCs in PAS-2 also were similar. CONCLUSION: Transfusion of PCs in PAS-2 significantly reduces the incidence of reactions. The 1-hour and 20-hour CCIs after transfusion of PCs in PAS-2 were significantly lower than the CCIs after transfusion of PCs in plasma. Because storage conditions of both PCs were found to be optimal, the decrease in CCIs after transfusion of PCs prepared in PAS-2 may be caused by rapid elimination of a subpopulation of P-selectin-positive platelets from the circulation.

Differences in residual white blood cell subset counts in buffy coat-depleted red cell concentrates prepared with bottom and top or quadruple-bag systems.

BACKGROUND: For preparation of buffy coat-depleted red cell concentrates (RCCs) in additive solution whole blood is currently collected in The Netherlands both in quadruple-bag and bottom and top bag systems. By using the quadruple-bag system both plasma and buffy coat cells are transferred into integrated satellite bags while the red cells remain in the collection bag. When bottom and top bags are used, the buffy coat remains in the collection bag while both red cells and plasma are transferred into satellite bags. The difference in processing prompted us to perform quantitative analysis of residual WBC subsets in buffy coat-depleted RCCs. Differences in removal of specific cells with the buffy coat could improve the outcome of additional filtration procedures aiming at complete removal of specific WBC subsets. STUDY DESIGN AND METHODS: The buffy coat was removed in semiautomated procedures (Optipress I; Compomat G4) from units of whole blood collected in both bag systems. Paired samples were taken before and after removal of the buffy coat for counting and analyzing WBC subsets by flow cytometry using subset-specific monoclonal antibodies. RESULTS: All RCCs met the criteria from the guidelines of the Council of Europe. The percentage of residual total WBCs was lower (p<0.001) in RCCs processed in bottom and top bag systems (26% Compomat and 18% Optipress) as compared to RCCs processed in quadruple-bag systems (43% Compomat). WBC subset analysis in RCCs processed in quadruple-bag systems showed approximately 25% of residual T cells, B cells and monocytes and 60% of residual granulocytes. In contrast, WBC subset analysis in RCCs processed in bottom and top bag systems showed approximately 2% residual T cells, B cells, and monocytes and 35% residual granulocytes; in about 45% of units, lymphocytes and monocytes were even below the detection limit of flow cytometry analysis. CONCLUSION: Buffy coat-depleted RCCs are currently processed in bottom and top bag or quadruple-bag systems, the former being superior to the latter due to selective depletion of lymphocytes and monocytes by 98% (2 logs). The bottom and top procedure is an evident contribution to leukodepletion in blood transfusion, both with and without additional filtration.

Platelets stored in a new-generation container differences between plasma and platelet additive solution II.

BACKGROUND AND OBJECTIVES: In order to preserve platelet concentrates (PCs) with high yields, a new polyolefin platelet storage container (PL 2410, 1.3L, Baxter, La Châtre, France) with increased gas permeability in combination with a larger surface area has been developed. The storage capacity was studied with platelets in plasma and platelets additive solution. MATERIALS AND METHODS: Platelet concentrate pools (PCs) of different yields suspended in either plasma (PCs-PL; n = 30) or PAS II (PCs-PAS; n = 37) were prepared. For preparation of PCs with a low, intermediate and high number of platelets 3, 5 and 6 buffy coat (BCs) were pooled with different volumes of plasma and 5 and 6 BCs were pooled with different volumes of PAS II, in order to obtain PCs of equal volumes comparable with routine conditions. All PCs were stored on a flatbed shaker at 22+/-2 degrees C and evaluated on days 1, 2, 5 and 7 by determining platelet and white cell counts, pH (37 degrees C), pO2, pCO2 and swirling score. RESULTS: Platelet yields ranged from 1.5 up to 5.5 x 10(11) platelets/U. On day 7 all PCs-PL (n = 4) with platelet yields above 4.5 x 10(11)/U had a pH value below 6.8 (range 5.91-6.79). While 7 of 8 PCs-PAS units with platelet yields above 4. 0 x 10(11)/U showed a pH value below 6.8 (range 6.31-6.70). CONCLUSION: During 7 days of storage in a new 1.3-liter platelet container, the pH was maintained above 6.8 in PCs in plasma with a yield between 1.5- and 4.5 x 10(11)/U; when PAS II was used, the maximum platelet yield allowed for comparable pH maintenance was somewhat lower (4.0 x 10(11)/U).

Improvement of platelet storage conditions by using new polyolefin containers.

BACKGROUND: Storage of pooled platelet concentrates (PCs) with yields above 3.0 x 10(11) platelets per unit in a 1-L PL-732 polyolefin container for 5 days often results in a drop in pH to below 6.0. Recently, new oxygen-permeable platelet containers (1-L PL-2410, 1-L and 1.5-L Compoflex) have been developed. The maximal platelet storage capacities of the new containers and the PL-732 were compared. STUDY DESIGN AND METHODS: Large platelet pools (n = 27) with platelet concentrations between 1.2 and 1.4 x 10(11) per L were made from 3 to 5 PCs prepared from buffy coats. The pools were divided in equal volumes among the PL-732 and the three new platelet containers. Platelet counts in the PCs ranged from 1.0 to 5.0 x 10(11) per unit. All PCs were stored on a flatbed shaker at 22 +/- 2 degrees C and evaluated on Days 1, 3, 5, and 7 by measuring platelet count, pH, pO2, pCO2, HCO3-, glucose, lactate, platelet swirling, and soluble p-selectin. RESULTS: Day 7 storage of PCs (n = 6) with yields between 3.0 and 4.0 x 10(11) platelets in PL-732 showed mean +/- SD pH values of 5.93 +/- 0.05 and lactate values of 32.3 +/- 7.9 mmol per L; in 4 of these 6 PCs, pH was below 6.0. In contrast, storage of these PCs in 1-L PL-2410 and 1.5-L Compoflex containers and of 2 of these 6 PCs in 1-L Compoflex containers showed pH values above 6.8. Lactate values were 15.5 +/- 1.3, 15.3 +/- 1.8, and 19.5 +/- 4.7 mmol per L, respectively (p < 0.001 vs. PL-732). The platelet storage capacity of the new containers with platelet yields between 4.0 and 5.0 x 10(11) per unit (n = 6) was evaluated. Day 7 storage of these PCs in the 1.5-L Compoflex showed an average pH value of 6.74 +/- 0.20; in 2 of 6 PCs, pH was below 6.8. The average pH value in the PL-2410 was 6.38 +/- 0.31, and in all PCs, pH was below 6.8. Average lactate values were 17.8 +/- 5.7 and 25.8 +/- 5.6 mmol per L (p < 0.05), respectively. Soluble p-selectin values on Day 7 of storage increased approximately twofold in all PCs. CONCLUSION: The new oxygen-permeable containers showed platelet quality comparable to that with the PL-732 and for longer storage periods and at higher platelet counts.

Evaluation of storage conditions of platelet concentrates prepared from pooled buffy coats.

Five days storage of pooled platelet concentrates (PCs) with high yields often results in a pH fall and poor platelet morphology despite the use of specific containers. In this study we evaluated two techniques for prolonged storage of PCs with high platelet counts, by measuring pH and platelet swirl. In routine procedures, 90 PCs, prepared from five buffy coats, were stored in a single 1-litre PL 732 container. After 5 days storage, 51 PCs with platelet counts below 3.1 x 10(11)/U showed a pH above 6.8 and optimal swirling scores. 29 of 39 PCs (74%) with yields above 3.1 x 10(11)/U showed pH values below 6.8 and poor swirling scores. These poor results can be prevented by shortening the duration of storage. PCs with high yields can be stored for 5 days either by dividing the concentrate into two containers or by adjusting the platelet count to 3.1 x 10(11)/U. In the latter procedure, we reduced the volume of the concentrate and found that of 102 PCs, only 9 showed poor values for pH and platelet swirl. The data clearly indicate the importance of measuring the prestorage platelet count to select the optimal procedure for concentrate storage.

Neutrophil granulation stained with May Grunwald Giemsa or pure Azure B-eosin Y quantified by image analysis.

Neutrophil granulation was quantified after staining with May Grunwald Giemsa or with the pure dyes Azure B and eosin Y. Spinner slides of the buffy coat of 3 normal subjects and 14 persons with different grades of toxic granulation were studied. Morphometric parameters were measured using an image analysis computer (Texture Analysis System, Leitz Wetzlar, FRG). The parameters for granulation varied over a wider range in Azure B (AzB) than in May Grunwald Giemsa (MGG) stained granulocytes. This is in accordance with the visual microscopy observation, that granulation is more pronounced after staining with AzB than MGG. The predominant shade of the nucleus was similar in both stains, whereas considerable and variable differences in the shade of the cytoplasm were found.

Vindesine, cisplatin, and bleomycin combination chemotherapy in non-small cell lung cancer: survival and quality of life.

Although combination chemotherapy is applied on a large scale in advanced non-small cell lung cancer (NSCLC), we still lack evidence indicating in which subsets of patients survival or quality of life might be improved. We studied these issues among a sample of 28 NSCLC patients with a high performance status, who received a tri-weekly vindesine, cisplatin, and bleomycin combination. Treatment was extended for an additional two courses only if a response was observed after the initial three courses. An overall response rate of 13/27 evaluable patients (48%) was found (complete response 1/27 and partial response 12/27) with a median response duration of 24 weeks. Median survival was 33 weeks (47 for responders and 26 for non-responders). Toxicity was primarily related to cisplatin, including severe nausea and vomiting and nephrotoxicity in 68% and 21% of the patients, respectively. Performance status and body weight dropped significantly during chemotherapy both among responders and non-responders. Performance status after discontinuation of chemotherapy approached pre-treatment scores in responders only. While the antitumor effect of this drug combination was confirmed, we conclude that treatment-associated toxicity and deterioration of the patients' well-being offset any potential survival advantage for the majority of patients.

DNA measurements in thin sections of lymphomas.

The DNA content of the nuclei of lymphomas and a reactive lymph node was studied by light absorption measurements in Feulgen-stained thin (2 microns) sections of lymph node biopsies, using the TAS image analysis system. For each nucleus, the integrated light absorbance at 548 nm wavelength was multiplied by the square root of the nuclear area to obtain a parameter for DNA independent from the nuclear size. In a hyperplastic reactive lymph node with follicular hyperplasia, the distribution of DNA X square root area was different in centrocytes and centroblasts, being compatible with a diploid centrocyte fraction and a mainly tetraploid centroblast fraction, in which hypertetraploid (octoploid?) nuclei were present. Similar measurements in three follicular centroblastic-centrocytic lymphomas gave similar results, with the centroblasts mainly tetraploid. The DNA distribution was studied in sections of lymphomas of high and low grades of malignancy, classified visually on the number of mitoses seen per field. The distribution width, from the smoothed histogram as the DNA X square root area values beyond the maximum peak frequency, appeared to be larger in the high-grade malignant lymphomas, in accordance with their higher number of mitoses.

A method to estimate the DNA content of whole nuclei from measurements made on thin tissue sections.

A microdensitometry method that allows estimation of the distribution of the DNA content of nuclei in thin tissue sections is described. The method is based on a theoretical model of Feulgen-stained spherical nuclei, of different sizes, in each of which the DNA is present as a homogeneous solution. In thin sections of nuclei of different sizes, the fraction of DNA per section is inversely proportional to the radius of the nucleus. Histograms of the product of DNA content and radius per nuclear section are independent of nuclear size but depend on total DNA content. The distribution of the total DNA content of nuclei in a section can be estimated from such a histogram. The results of the measurements of a Feulgen-stained rat liver section are described.

Stability of azure B-eosin Y staining solutions.

The stability of azure B-eosin Y staining solutions of varying composition and of a routine May Grunwald Giemsa (MGG) stain were studied by analysis of the density histogram of white blood cells obtained by an image analysis computer. The stability appeared to be variable and depended on the concentration of the dyes, the molarity of the buffer solutions and the presence of dimethylsulfoxide (DMSO) as a stabilizer. Although most staining solutions including the routine MGG stain showed marked loss of staining capacity soon after preparation, it was possible to obtain an azure B-eosin Y mixture with very satisfactory staining properties which did not decrease during 8 h after its preparation.

Prevalence of achlorhydria in a normal population and its relation to serum gastrin.

In a middle-aged group of normal subjects achlorhydria after maximal stimulation with pentagastrin was found in 12 out of 366 males (3%) and 2 out of 198 females (1%); 13 of these 14 (93%) had an elevated serum gastrin, of which nine were more than two times normal. In another 11 persons a moderately elevated serum gastrin - less than two times normal - was found in absence of achlorhydria. At least seven of them had hypochlorhydria. Of 540 persons with a normal serum gastrin one had achlorhydria. Determination of the serum gastrin appeared to be fairly reliable as a method of detecting achlorhydria.

Krypton-81m ventilation studies as a parameter for lung capacity after lobectomy.

Spirometry and krypton-81m lung scans were performed in 17 patients before lobectomy was carried out for bronchogenic carcinoma to determine whether post-operative spirometric values could have been predicted from the pre-operative data. The patients were followed up at 3, 6 and 12 months post-operatively for lung function testing. It emerged from this study that good post-operative spirometric values could be predicted for VC (r = 0.80), and for FEV1 (r = 0.88), and to a much lesser degree for the TLC (r = 0.51) at 6 months after surgery. With time TLC and VC tended to increase, whilst the FEV1 remained at the same level.

Parietal cell antibodies in relation to basal serum gastrin in a normal population.

Parietal cell antibodies, basal gastric pH and serum gastrin were determined in 544 normal non-achlorhydric subjects. Serum gastrin was significantly higher in subjects with parietal cell antibodies, both in the group with a weakly positive and in that with a strongly positive fluorescence score. The elevated serum gastrin level in subjects with circulating parietal cell antibodies was independent of gastric pH. It is suggested that parietal cell antibodies may interfere with gastrin receptors on the membrane of the parietal cell, thus influencing the feedback mechanism between the secretion of acid and gastrin, and resulting in an increase of the basal serum gastrin level at an unchanged basal gastric pH.

Classification of immature and mature cells of the neutrophil series using morphometrical parameters.

Quantitative morphological data of six classes of immature and mature cells of the neutrophil series of the bone marrow of normal persons were used for statistical classification experiments (myeloblasts, promyelocytes, myelocytes, metamyelocytes, bands and segments). On each cell, parameters were measured directly from the image or calculated from the shape of the density histogram or the counting densitogram using a Texture Analysis System (E. Leitz, Wetzlar, Germany). The parameters were analyzed with the interactive statistical pattern recognition system ISPAHAN. One half of the data were used as a learning set and the other half as the test set. The parameters were compared according to their performance in discrimination between the classes, alone and in combinations. Parameters not contributing to an improvement of the discrimination were disregarded. Eleven parameters were selected and used for classification by two different methods: a stepwise and a "one-shot" method. Stepwise classification resulted in a 79% correct classification rate. Most errors occurred between cell classes in neighboring stages of maturation. In 96% of all cases the computer classification was either in accordance with that of the technician or with a cell class of a neighboring maturation stage. One step classification by the computer was in agreement with the technicians in 82% of the cases. For 98% of the cells the computer classification was either in accordance with that of the technician or with a cell class of a neighboring maturation stage. The data set was collected by two technicians, operating independently. Differences in their interpretation of the maturation stage were found by comparing the performance of classifiers based on both cell samples. Since the images of the cells were not available for reexamination, the causes of disagreement in classification between the technicians and between computer and technicians could not be evaluated.

Is discrimination between type A and B atrophic gastritis clinically useful in achlorhydria?

We tested the validity of the concept that chronic atrophic gastritis can be subdivided into type A and B in hospital patients and normal subjects with proven pentagastrin-refractory achlorhydria. Classification was based on the determination of the basal serum gastrin and parietal cell antibodies. Of 59 hospital patients with achlorhydria, 71% could be classified as belonging to either type A or B; for 29% the criteria for neither type were fulfilled. Of 14 asymptomatic persons with achlorhydria found in 564 normal persons, five could be classified as having type A gastritis, and one as type B gastritis. In eight (53%) persons, an elevated serum gastrin was found in the absence of parietal cell antibodies, representing an intermediate type of atrophic gastritis. Because one-third of the hospital patients and more than half the persons with achlorhydria in a normal population had to be classified as belonging to an intermediate type, the discrimination between type A and B atrophic gastritis in achlorhydria seems to be of limited practical value.