Pseudobasophilia as an erroneous white blood cell differential count with a discrepancy between automated cell counters: report of two cases

We report two cases that showed erroneous white blood cell differential counts by automated cell counters. Each case showed an interesting discrepancy of differential count between cell counters, and marked pseudobasophilia was observed by one of the two counters. The first patient was a 44-year-old female who suffered from multiple myeloma for more than one and a half years. Increased myeloma cells (43%) in peripheral blood were counted as basophils by the ADVIA 120, and as monocytes by SE-9000, respectively. The second patient was a 72-year-old female diagnosed as having chronic myelomonocytic leukemia. Dysgranulopoietic neutrophils (50%) and monocytes (31%) were increased in the peripheral blood. Dysgranulopoietic neutrophils were counted as basophils by STKS. In contrast, about half of the increased monocytes were counted as neutrophils by the ADVIA 120. These interesting findings highlight the importance of microscopic examination of the blood film in routine laboratory practice, and automated cell counters, especially for the hematologic patients, cannot completely substitute for it. These results also imply that at least some subpopulations with different membrane or cytoplasmic properties may exist even in the similarly classified cells.     

PKCdelta associates with and is involved in the phosphorylation of RasGRP3 in response to phorbol esters

RasGRP is a family of guanine nucleotide exchange factors that activate small GTPases and contain a C1 domain similar to the one present in protein kinase C (PKC). In this study, we examined the interaction of RasGRP3 and PKC in response to the phorbol ester PMA. In Chinese hamster ovary or LN-229 cells heterologously expressing RasGRP3, phorbol 12-myristate 13-acetate (PMA) induced translocation of RasGRP3 to the perinuclear region and a decrease in the electrophoretic mobility of RasGRP3. The mobility shift was associated with phosphorylation of RasGRP3 on serine residues and seemed to be PKCdelta-dependent because it was blocked by the PKCdelta inhibitor rottlerin as well as by a PKCdelta kinase-dead mutant. Using coimmunoprecipitation, we found that PMA induced the physical association of RasGRP3 with PKCdelta and, using in situ methods, we showed colocalization of PKCdelta and RasGRP3 in the perinuclear region. PKCdelta phosphorylated RasGRP3 in vitro. Previous studies suggest that ectopic expression of RasGRP3 increases activation of Erk1/2. We found that overexpression of either PKCdelta or RasGRP3 increased the activation of Erk1/2 by PMA. In contrast, coexpression of PKCdelta and RasGRP3 yielded a level of phosphorylation of Erk1/2 similar to that of control vector cells. Our results suggest that PKCdelta may act as an upstream kinase associating with and phosphorylating RasGRP3 in response to PMA. The interaction between RasGRP3 and PKCdelta points to the existence of complex cross-talk between various members of the phorbol ester receptors which can have important impact on major signal transduction pathways and cellular processes induced by phorbol esters or DAG     

Mineralogical and chemical changes in pyrite after traditional processing for use in medicines

Pyrite has been the most commonly used medicinal mineral, and its toxicity was reduced by traditional processing operations including heating and quenching in vinegar. To verify the scientific effects of this process, pyrite was processed at temperatures up to 850 degrees C and through as many as five processing cycles. A metal extraction test was carried out from the processed pyrites on the assumption that pyrite medicines with the lowest toxic metal content would be most desirable. Increasing temperature and the number of processing cycles promoted phase change of pyrite to hematite, reduction of toxic metals in pyrite and their concentrations in the extraction solutions. However, the relationships between variations in extracted elements and the number of processing cycles at the same processing temperature were not clearly defined. Heating temperature is more important than the number of processing cycles for effective processing, and pyrite should be processed at the highest possible temperature in order to diminish highly toxic metals such as As and Pb.     

Intestinal parasites in children with neoplasms

Stool specimens taken from 50 children with malignancy and from 92 healthy children were investigated for intestinal parasites, using the modified formol ethyl acetate concentration method, and native-lugol, trichrome and Kinyoun acid-fast stain methods. Thirty-eight (76.0%) of the 50 patients had lymphoma or leukemia and were considered immunosuppressed. Several different parasites were found in 21 (42.0%) of the 50 patients with malignancy and in 16 (47.3%) of the 38 patients with immune deficiency compared to in only 16 (17.3%) of the 92 healthy children. The incidence of parasites in patients with malignancy or immunosuppression was significantly higher than in the healthy control group (p<0.01, p<0.01).     

Matrix metalloproteinase-2, membranous type 1 matrix metalloproteinase,and tissue inhibitor of metalloproteinase-2 expression in ectopic and eutopic endometrium

OBJECTIVE: To investigate expression of matrix metalloproteinase-2 (MMP-2), membranous type 1 matrix metalloproteinase (MT1-MMP), and tissue inhibitor of metalloproteinase-2 (TIMP-2) in ectopic and eutopic endometrium from women with and without endometriosis throughout the menstrual cycle. DESIGN: Molecular studies in human tissue. SETTING: Reproductive immunology laboratory of a university medical center. PATIENT(S): Fifty-three premenopausal woman (23 with endometriosis and 30 without endometriosis) undergoing laparoscopic surgery. Endometrium and ectopic endometriosis tissue were obtained at the time of surgery. MAIN OUTCOME MEASURE(S): Messenger RNA and protein expression from eutopic and ectopic endometrium was analyzed by using quantitative competitive polymerase chain reaction, zymography, and Western blot assay. RESULT(S): Uterine endometrium from women with endometriosis expressed higher levels of MMP-2 and MT1-MMP and lower levels of TIMP-2 than did endometrium from normal women. CONCLUSION(S): Eutopic endometrium from patients with endometriosis may be more invasive and prone to peritoneal implantation because of greater expression of MMP-2 and MT1-MMP and lower expression of TIMP-2 messenger RNA, compared with endometrium from women without endometriosis. Thus, increased proteolytic activity may help to explain the invasive factors that result in endometriosis.     

Identification of a compound that directly stimulates phospholipase C activity

Phosphoinositide-specific phospholipase C (PLC) plays a pivotal role in the signal transduction of various cellular responses. However, although it is undeniably important that modulators of PLC activity be identified, no direct PLC activity modulator has been identified until now. In this study, by screening more than 10,000 different compounds in human neutrophils, we identified a compound that strongly enhances superoxide-generating activity, which is well known to be PLC-dependent. The active compound 2,4,6-trimethyl-N-(meta-3-trifluoromethyl-phenyl)-benzenesulfonamide (m-3M3FBS) stimulated a transient intracellular calcium concentration ([Ca(2+)](i)) increase in neutrophils. Moreover, m-3M3FBS stimulated the formation of inositol phosphates in U937 cells, indicating that it stimulates PLC activity. The compound showed no cell-type specificity in terms of [Ca(2+)](i) increase in the various cell lines including leukocytes, fibroblasts, and neuronal cells. We also ruled out the possible involvement of heterotrimeric G proteins in m-3M3FBS-stimulated signaling by confirming the following: 1) pertussis toxin does not inhibit m-3M3FBS-induced [Ca(2+)](i) increase; 2) m-3M3FBS does not stimulate cyclic AMP generation; and 3) the inhibition of G(q) by the regulator of G protein-signaling 2 does not affect the m-3M3FBS-induced [Ca(2+)](i) increase. We also observed that m-3M3FBS stimulated PLC activity in vitro. The purified isoforms of PLC that were tested (i.e., beta2, beta3, gamma1, gamma2, and delta1) were activated by m-3M3FBS and showed no isoform specificity. Taken together, these results demonstrate that m-3M3FBS modulates neutrophil functions by directly activating PLC. Because m-3M3FBS is the first compound known to directly activate PLC, it should prove useful in the study of the basic molecular mechanisms of PLC activation and PLC-mediated cell signaling.     

Abnormalities of erythrocyte membrane proteins in Korean patients with hereditary spherocytosis

Hereditary spherocytosis (HS) is a common inherited erythrocyte membrane disorder characterized by chronic hemolytic anemia. Clinical manifestations and biochemical abnormalities of HS are heterogeneous. In this study, we investigated erythrocyte membrane protein defects in 27 Korean HS cases. Utilizing both the Fairbanks system and the Laemmli system, sodium dodecyl sulfate polyacrylamide gel electrophoresis of erythrocyte membrane proteins was performed. Proteins were stained with Coomassie brilliant blue and gels were scanned using a densitometer. We detected spectrin deficiency in 7.4% of cases (2/27), ankyrin deficiency in 29.6% (8/27), combined spectrin and ankyrin deficiency in 3.7% (1/27), band 3 deficiency in 11.1% (3/27) and protein 4.2 deficiency in 14.8% (4/27). Membrane protein deficiencies were not observed in nine cases (33.3%, 9/27). Members of two of seven families tested showed the same protein defects as the proband. Ankyrin deficiency alone and combined with spectrin deficiency accounted for 33.3% of cases (9/27), and they were the most common biochemical defects in Korean HS cases. Protein 4.2 deficiency caused HS more frequently in Koreans than in Caucasians.     

In vivo laxity of stable versus anterior cruciate liagment-injured knees using a navigation system: a comparative study

We compared antero-posterior translation and internal–external rotation of the tibia in stable knees without anterior cruciate ligament (ACL) injury with those of ACL injured knees using a navigation system and suggest an objective data. Forty-four patients treated for a meniscal tear without ACL injury were allocated to stable group, and 41 patients were allocated to ACL injury group. Antero-posterior displacement and rotation of knees were measured in 0, 30, 60 and 90 degrees of flexion using navigation. Mean anterior displacements were 3.6 ± 2.0, 6.7 ± 2.7, 6.0 ± 2.4 and 4.7 ± 1.8 mm at 0, 30, 60 and 90 degrees of flexion, respectively, in stable group, and 6.8 ± 3.6, 14.7 ± 3.5, 11.9 ± 4.6 and 8.5 ± 4.0 mm in ACL injury group. Mean total rotation values were 18.8 ± 4.5°, 31.4 ± 4.2°, 30.1 ± 5.1° and 29.2 ± 5.9° in stable group and 22.7 ± 6.9°, 37.6 ± 5.8°, 34.0 ± 9.4° and 31.8 ± 8.8° in ACL injury group. Quantitative values of antero-posterior translations and rotations of stable and ACL injured knees were obtained using a navigation system. The laxity data may be useful to establish the diagnosis of an ACL injury and evaluation of post-operative results.     

Self-splicing activity of the mitochondrial group-I introns from Aspergillus nidulans and related introns from other species

The mitochondrial genome of Aspergillus nidulans contains several group-I introns. Each one has been assayed for its ability to self-splice in vitro in the absence of proteins. The intron from the apocytochrome b gene is unusual among subgroup IB4 introns in being able to self-splice, unlike a similar intron from Saccharomyces cerevisiae. The first intron in the cytochrome oxidase subunit-1 gene self-splices but only correctly completes the first step of splicing; cryptic 3′ splice-sites are recognized instead and these are also used at a low frequency in vivo. The highly homologous intron from Podospora anserina completes both steps in vitro. The remaining introns do not self-splice. The correlation between subgroup category, the likely presence of specific tertiary interactions, and self-splicing activity is discussed. Received: 16 May / 25 August 1997