Biomedical subjects
Publications and source records attributed to G Sirchia.
Donor organ procurement is a world-wide problem. In Italy it is particularly so and the reasons for this are investigated. An overall increase in the number of donors has been noted in 1994 and the first 8 months of 1995, and ways of continuing this encouraging trend should be pursued by improvements in education, legislation, and hospital organization.
From dialyzable leucocyte extracts (DLE) we have purified a hydrophilic low-mol. wt. factor (about 1 kDa) which we have named lymphocytic suppressor factor (LSF) as it is able to suppress antigen- and mitogen-induced lymphocyte transformation and to prolong allograft survival in C57b/6N mice (H-2b) transplanted with fully mismatched skin from C3H/HeN mice (H-2k). At the molecular level LSF acts by inhibiting DNA replicational and transcriptional processes in activated lymphocytes, isolated rat hepatocyte nuclei, and cell-free systems. Amino acid analysis indicates that LSF is a peptide composed of Asp, Glu, Ser, Thr, Ala, Gly, Arg and probably Met, with the N-terminus blocked, possibly by pyroglutamic acid. When combined "in vitro" with cyclosporine A (CsA), LSF increased about 20 times the potency of CsA in inducing suppression of mitogen-stimulated lymphocytes. In C57b/6N mice with skin graft from C3H/HeN mice and undergoing immunosuppression with CsA (50 mg/kg/day), the splenocyte LSF content increased about 5 times. However, LSF values returned to normal in mice recovering normal responsiveness due to progressive withdrawal of CsA. These data show that LSF has an important role in the development and maintenance of CsA-induced immunosuppression. We suggest that, by influencing DNA replicational and transcriptional processes of lymphocytes, LSF may play a role also in the onset and progression of AIDS induced by retroviruses.
We characterized serologically 5 anti-C (4 IgM and 1 IgG), 3 anti-c (2 IgM and 1 IgG), 4 anti-E (1 IgM and 3 IgG), 4 anti-e (3 IgM and 1 IgG) and 46 anti-D (16 IgM and 30 IgG) monoclonal antibodies, provided by the Rh Section of the Third International Workshop and Symposium on Monoclonal Antibodies against Human Red Blood Cells and Related Antigens (1996) for their ability to detect weak and variant antigens. The agglutination patterns were established using untreated and papain-treated red blood cells in a column agglutination technology system (BioVue, Ortho). Significant differences were found between the IgM and IgG antibodies. The papain treatment seemed to be important for IgM but not for IgG antibodies. Almost all of the IgM anti-D antibodies detected untreated DIV samples and almost all of the IgG anti-D antibodies detected untreated weak D samples. Both IgM and IgG anti-D antibodies showed the highest number of negative reactions with DVI and Rh 33 red blood cells. The CwCw sample was detected by only one of the 4 anti-C IgM MAbs using enzyme-treated red blood cells. All anti-c MAbs were able to detect treated Cx samples. Because of the small number of weakly expressed E and e samples, definitive conclusions cannot be drawn on the ability of these antibodies to detect these antigens.
Guidelines, algorithms and recommendations have been issued in the attempt to ensure appropriateness of transfusion practice, but the results are less than satisfactory, mainly due to the difficulty to turn paper procedures into actual practice. In our hospital we have tried to overcome this difficulty through the implementation of a quality assurance programme which includes giving the privilege of nonurgent blood prescription to a limited number of physicians and a computerized prospective audit of blood requests. The latter is performed through verification of the compliance of blood requests, which are designed to include a patient's laboratory and clinical data, with hospital guidelines for the proper use of blood. In the 12 months since implementation of the computerized prospective audit the transfusion service has evaluated 7884 requests. Of these, 63.4% (n = 4998) were for red blood cells, 21.1% (n = 1664) for platelets and 15.5% (n = 1222) for fresh frozen plasma. The prospective audit showed that 96.8% and 98.1% of requests for red units and platelets were appropriate, respectively. Conversely, approximately 27% of plasma requests did not comply with guidelines, mainly because the evidence of coagulopathy was missing. However, inappropriateness of plasma requests for elective general surgery decreased from 39% at the onset of the programme to 14% in the last trimester considered. Moreover, the evaluation by retrospective audit of the proportion of patients transfused with both red blood cells and plasma in the perioperative period out of those transfused with red blood cells only, as an indicator of unwanted reconstitution of whole blood, showed that this proportion decreased from 47.6% (320/672) in the 12 months before implementation of computerized audit to 37.8% (244/646) in the following 12 months (difference = -9.8%, 95% confidence interval of the difference from -4.5% to -15.1%; P < 0.005 by chi 2 test). Our initial experience, together with the present system, shows that (1) the restriction of nonurgent blood prescription to a group of clinicians more educated in transfusion medicine than average clinicians practicing in a large multispecialty hospital is feasible; (2) prospective audit is a useful tool for assuring the quality of blood requesting.
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BACKGROUND: A quality control program of white cell (WBC) reduction in red cells at the bedside was implemented, based on postfiltration counting in a Nageotte chamber of the residual WBCs from samples taken from a segment of the transfusion set, after 1-in-10 sample dilution with Türks's solution. During a 1-year quality control program, 5.1 percent of counted units had apparent filtration failures, that is, WBC counts exceeding 5 x 10(6) per unit. The cause(s) for these apparent failures were investigated. STUDY DESIGN AND METHODS: In Study 1, residual WBCs from 150 buffy coat-free red cells filtered through one type of filter at 4 degrees C, 20 to 24 degrees C, or 27 degrees C in 5 to 10 minutes, 50 to 100 minutes, or 100 to 200 minutes were counted as described above. In Study 2, residual WBCs in samples collected from segments of the transfusion set and from the postfiltration bags were counted in parallel by a new, more sensitive counting method. In this method, 5 mL of filtered red cells was diluted with 20 mL of 3-percent paraformaldehyde and centrifuged, the pellet was resuspended to 500 microL with a lysis solution, and the WBCs were counted in a Nageotte chamber. In Study 3, residual WBCs were counted by the 3-percent paraformaldehyde method in samples from postfiltration bags of 1- to 2-day-old buffy coat-rich red cell units filtered through a second type of filter. Filtration was started within 30 minutes of the removal of the unit from the refrigerator, ambient temperature was 20 to 24 degrees C, and the median filtration time was 90 minutes per unit. RESULTS: Study 1: Median WBC counts per unit increased progressively from 51,000 at 4 degrees C to 934,000 at 27 degrees C, with intermediate values at 20 to 24 degrees C. In no unit did the WBC count exceed 5 x 10(6) if filtration at 20 to 24 degrees C was completed within 100 minutes, while counts in excess of 50 x 10(6) were found at 20 to 24 degrees C and at 27 degrees C with filtration times of 100 to 200 minutes, and 50 to 100 minutes, respectively. Study 2: The relation between segment and postfiltration bag WBC counts obtained by the 3-percent paraformaldehyde method was poor, with the latter being almost always lower than the former. Study 3: None of the 120 units filtered through the second type of filter at 20 to 24 degrees C in 50 to 100 minutes contained more than 3.2 x 10(6) WBCs; the median value was 147,000 WBCs per unit. CONCLUSION: On the basis of the results with the 3-percent paraformaldehyde method, which showed the unreliability of segment counts, a new policy was adopted for quality control of bedside WBC reduction, based on controlling the time of and temperature at transfusion. Bedside WBC reduction in 1- to 2-day-old red cells performed with the second type of filter at 20 to 24 degrees C in less than 100 minutes per unit allowed the preparation of units that meet the standard of fewer than 5 x 10(6) WBCs in all tested cases. Bedside WBC reduction with the second type of filter and under the controlled conditions reported seems effective.
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BACKGROUND & AIMS: The association of liver disease with hepatitis C virus (HCV) genotypes mainly refers to patients with serious liver damage; little information is available on symptomless carriers. The aim of this study was to investigate the correlation of genotypes with clinical course, risk factors for infection, and antibody to HCV reactivity in asymptomatic subjects. METHODS: One hundred nine viremic blood donors with at least 1 year of follow-up were studied; 41 underwent liver biopsy. Genotypes were determined by line-probe assay. RESULTS: Genotype 1 was found in 47 (43.1%), genotype 2 in 48 (44%), genotype 3 in 8 (7.3%), genotype 4 in 2 (1.8%), and coinfections in 4 (3.7%). The relative risk (RR) for a raised pattern of alanine aminotransferase, aspartate aminotransferase, and gamma-glutamyltranspeptidase was 2.1 (confidence interval [CI], 1.4-3.2), 1.7 (CI, 1.2-2.4), and 2.8 (CI, 1.6-4.9) in subjects with genotype 1 vs. 0.4 (CI, 0.2-0.7), 0.4 (CI, 0.3-0.7), and 0.4 (CI, 0.2-0.8) in subjects with genotype 2. Chronic hepatitis was found in 68%; the RR of chronic hepatitis was similar for genotypes 1 and 2 (RR, 1.1 [CI, 0.8-1.7] vs. RR, 1.0 [CI, 0.7-1.6]). Reactivity to NS4-derived antigens was infrequent in type 2-infected subjects. CONCLUSIONS: Genotype 2 was as frequent as genotype 1 but associated with less liver function impairment. The high prevalence of chronic hepatitis should be considered in counseling viremic asymptomatic donors.
We describe the activities of the Milan Cord Blood (CB) Bank and of the Italian Cord Blood Network. By October 31, 1995, 763 units were banked in Milan. Of these, 8 units were used to perform 4 related and 4 unrelated transplants in 5 children and 3 adult patients. Early cord clamping after delivery was found to be crucial to increase the volume of CB collected. This procedure does not seem to be detrimental to the newborn. Of various red cell sedimenting agents used to reduce the unit volume and concentrate progenitor cells, 3% gelatin seems to be associated with the best yields. After a preliminary experience of 2 years, the Italian Cord Blood Network (Gruppo Italiano Amplificazione Cellule Emopoietiche, GRACE) was founded in 1995. The initial activities of GRACE are aimed at the development at the national level of CB banking standard operative procedures in agreement with the draft issued by The North American Task Force for The Development of Standards for Hematopoietic Cell Transplantation. Moreover, a wet workshop has been organized to standardize colony-forming unit (CFU) evaluation. The main goal in Milan is the collection of 5000 CB units. Other issues of interest include CB volume reduction, hematopoietic progenitor purification, ex vivo expansion prior to transplantation, and experimental protocols for gene transfer, such as those related to the multidrug resistance (MDR) gene.
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Autologous- or allogeneic-bone marrow transplantation are increasingly used to overcome the myelosuppressive effects of high dose chemotherapy administered to cancer patients. Transfer of the multidrug resistance (MDR) gene in hemopoietic progenitors has been proposed as a tool to administer higher and possibly more curative doses of chemotherapy. Murine models have demonstrated that retrovirus-mediated MDR transfer in bone marrow cells can render animals resistant to myeloablative doses of Taxol, and in vitro studies have shown that MDR-transduced human CD34+ cells can generate drug-resistant multipotential hemopoietic progenitors such as long term culture-initiating cells. Given these results, phase I clinical trials are currently under way to evaluate feasibility and treatment-related toxicity of MDR gene transfer in cancer patients by means of safe retroviral vectors. Finally, Taxol treatment of MDR transduced mice and human CD34+ cells have indicated that MDR is a dominant selectable marker in vitro and in vivo, and vectors carrying both MDR and non selectable genes such as beta-globin or glucocerebrosidase could be used in the next future for gene therapy of inherited disorders like thalassemia or Gaucher disease.
Umbilical cord blood (CB) has been widely used for related and unrelated transplants in pediatric patients. We present the case of an adult with secondary AML who received an unrelated, one-antigen mismatched CB transplant due to the lack of a matched donor. The patient was a 26-year-old female (35 kg/bw) who had received an autologous bone marrow transplant for Hodgkin's disease in April 1994 and, 6 months later, developed secondary MDS (RAEB, 46, XX, -7, +mar), which slowly evolved into acute myelogenous leukemia. In May 1995, she was transplanted with a 165 ml CB unit containing a total of 1.6 x 10(9) nucleated cells, 11 x 10(6) CD34+ cells and 7.2 x 10(5) CFU-GM. GVHD prophylaxis consisted of standard CsA and methotrexate. Myeloid engraftment occurred on day +28 (PMN > 500) and full donor chimerism was confirmed twice (on days +33 and +56) by means of cytogenetics and DNA microsatellite analysis. Erythroid and megakaryocytic engraftment was documented by immunohistochemical analysis of a bone marrow biopsy on day +40, showing the presence of erythroblastic islands and isolated CD61+ immature cells. The patient did not develop GVHD but died on day +56 from idiopathic interstitial pneumonia and multiorgan failure. To our knowledge, this is one of the first case reports of unrelated mismatched CB transplantation in an adult.
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Matched related cord blood transplantation (CBT) has been successfully used to rescue patients undergoing myeloablative therapy. However, few data are available on the kinetics of hematological and immunological reconstitution of CBT recipients. We have investigated the hematological engraftment and immune recovery following related CBT in three patients, with acute lymphoblastic leukemia, aged 10, 9 and 7 years and with a body weight of 31, 40 and 25 kg, respectively. All patients engrafted and none experienced acute or chronic graft-versus-host disease. The time needed to achieve granulocyte recovery was 13, 26 and 29 days, respectively and platelet recovery occurred in 28, 49 and 51 days. All patients presented a marked increase of HbF, the values observed being much greater than those documented in patients given marrow transplantation and comparable with those observed in normal children in the first year of age. The recovery of T cell immunity, as well as that of natural killer subpopulations, mimicked that described in BMT recipients, a quicker return of CD8+ T cells determining the characteristic inversion of CD4/CD8 ratio. An impressive increase in the percentage and absolute number of B lymphocytes, apparently not related to viral infections, was demonstrable in all three cases. These data suggest that CBT recipients can experience a slight delay in hematological recovery when compared with patients given BMT. The reconstitution of erythropoiesis seems to recapitulate the ontogenetic pattern and the kinetics of recovery of the immune system reproduce that observed after BMT with the peculiarity of B cell expansion in peripheral blood.
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The introduction of genomic HLA-DR typing has stimulated a re-evaluation of the role of HLA-DR compatibility on cadaver kidney transplantation. We retrospectively studied the influence of HLA-DRB1 matching on the survival of 416 patients using univariate and Cox regression analysis as well as its influence on the occurrence of rejection episodes and on creatinine level at the 3rd month in the 198 recipients for whom these data were available. The following parameters were also considered: HLA-A,B compatibility, donor and recipient age, graft number, pre-transplant blood transfusions and panel reactive antibodies (PRA). Twenty-four month graft survival was 100% for transplants with zero mismatches (n = 47), 87.9% for those with one mismatch (n = 191) and 81.3% for those with two mismatches (n = 178). In the Cox model, HLA-DRB1 matching was the most significant variable influencing graft survival (47% of chi 2 P = 0.001), followed by HLA-A,B matching (23%, P = 0.02) and donor age (19%, P = 0.04). Ninety-two percent of the patients with zero mismatches experienced no rejection episodes in the first 3 posttransplant months compared with 62% and 41% of patients with one and two mismatches, respectively. Mean creatinine level (mg/dl) was 1.2, 1.4, and 1.5 in patients with zero, one, and two mismatches, respectively. Should these results be confirmed by prospective studies, HLA-DRB1 compatibility will have to be considered as an organ allocation criterion.