Evaluation of the analytical performances of a portable, 18-parameter hemometric system using capillary blood samples for blood donor enrolment.

BACKGROUND AND OBJECTIVES: Blood donor enrolment process is frequently based on the sole capillary haemoglobin (Hb) evaluation while platelet donors by apheresis also requires platelet (Plt) count. The 'sole Hb' approach prevents a complete donor evaluation and does not allow Plt donor enrolment. To extend blood counts before donations, we evaluated the performances of a multiparametric counter using capillary blood. MATERIALS AND METHODS: The ABX Micros 60 (Micros 60) blood analyzer was employed on capillary blood and compared with venous counts by a reference counter (Coulter AcT 5diff) in a first series of 416 donors and in a second series of 136, after a 3-month period of routine use of this study counter. An average of 50 microl of capillary blood was collected whose 10 microl had been aspirated by Micros 60. RESULTS: High correlations were found between capillary counts using Micros 60 and venous counts using the reference counter. Mean Plt counts differed of 37 x 10(9)/l less for capillary approach in the first series of comparisons, but decreased to 10 x 10(9)/l less in the second series due to a greater expertise of operators in capillary sampling. All other parameters were accurate and never reached clinical relevance albeit they showed statistically significant differences. CONCLUSION: Data on Micros 60 demonstrated that capillary predonation counts may represent a feasible and effective approach to realize an accurate enrolment process of blood and Plt donors.

Hepatitis B virus blood screening: impact of nucleic amplification technology testing implementation on identifying hepatitis B surface antigen non-reactive window period and chronic infections.

BACKGROUND: Despite improvements in hepatitis B surface antigen (HBsAg) test sensitivity, post-transfusion hepatitis B virus (HBV) infection still occurs because HBsAg is undetectable during the early window phase (WP) of the infection, in the convalescence core window phase of the infection, or in serologically silent chronic hepatitis or in mutant forms of HBV. HBV-DNA screening using high sensitivity nucleic amplification technology (NAT) assays has recently been introduced to reduce the residual risk of transmission of HBV by transfusion of blood components. MATERIALS: Over 1 year 75 063 donations were individually screened for HBV-DNA by the Ultrio Procleix assay on the Tigris platform. The donations were collected in the Latium region, an area of the central Italy, and they accounted for the 40% of the total blood units collected in this area per year. The initial reactive samples were re-tested and confirmed by the discriminatory HBV assay. Additional HBV serological markers were also performed. Suspected WP infections were followed-up to monitor the development of the immune response. All HBV-DNA-positive donors were called back to check up their infectious status. RESULTS: The results of testing the 75 063 donations are: 33 donations HBsAg positive, 31 out of them HBV-DNA-positive and two HBV-DNA negative; 22 donations HBsAg-negative but HBV-DNA positive with low viral load. Six of the 22 were found to be consistently HBV-DNA reactive whereas the remaining 16 donations showed inconsistent results on multiple NAT retesting. One WP infection was confirmed by the follow-up of the donor for 3 months following the index blood donation. CONCLUSIONS: In the donor population of the Latium region, NAT screening has revealed a higher than expected number of donors who were HBsAg non-reactive but HBV-DNA-positive with three donors showing HBV-DNA as the only marker of infection. The adoption of genome screening has increased the safety of the blood supply and has also contributed to the protection of donor health by identifying either WP or clinically silent infections.

Risk management of HBsAg or anti-HCV positive healthcare workers in hospital.

Recommendations are made for controlling the transmission of the hepatitis B and hepatitis C viruses from healthcare workers to patients. These recommendations were based both on the literature and on experts' opinions, obtained during a Consensus Conference. The quality of the published information and of the experts' opinions was classified into 6 levels, based on the source of the information. The recommendations can be summarised as follows: all healthcare workers must undergo hepatitis B virus vaccination and adopt the standard measures for infection control in hospitals; healthcare workers who directly perform invasive procedures must undergo serological testing and the evaluation of markers of viral infection. Those found to be positive for: 1) HBsAg and HBeAg, 2) HBsAg and hepatitis B virus DNA, or 3) anti-hepatitis C virus and hepatitis C virus RNA must abstain from directly performing invasive procedures; no other limitations in their activities are necessary. Infected healthcare workers are urged to inform their patients of their infectious status, although this is left to the discretion of the healthcare worker; whose privacy is guaranteed by law. If exposure to hepatitis B virus occurs, the healthcare worker must undergo prophylaxis with specific immunoglobulins, in addition to vaccination.

Elevated alanine aminotransferase in blood donors: role of different factors and multiple viral infections.

Many different aetiological agents stimulate alanine aminotransferase (ALT) production. Viral markers and other aetiologies were investigated in 2166 individuals, randomly selected from 10,000 consecutive blood donors. Elevation of ALT was found in 10.8% of subjects. Grouping donors according to ALT level and correlating with, respectively, hepatitis B core antibody (HBcAb), cytomegalovirus antibody alone, or associated with HBcAb, showed similar findings (high ALT 11.1%, normal 11.6%; high 85.4%, normal 81.4%; high 10.2%, normal 11.0%, respectively). Hepatitis C virus (HCV) antibody was found to be significantly associated with elevated ALT levels (high 1.7%, normal 0.26%). Other causes of ALT elevation were alcohol abuse (17%), obesity (25%) and dyslipidaemia (38%), but in 11% there was no obvious aetiology. Although HCV is a rare cause of elevated ALT in blood donors, it seems to be the only virus, among those tested, to account for liver damage. This may be due to the non-protective role of HCV antibody, the low specificity of ALT, or the pathogenic role of uninvestigated viruses.

Molecular analysis of HTLV-I and HTLV-II isolates from Italian blood donors, intravenous drug users and prisoners.

Using molecular methods three or five major variants of HTLV-I have been identified; moreover two subtypes of HTLV-II defined as HTLV-IIa and HTLV-IIb with six variants within each of these groups have been described. In the present study we analysed proviral DNA obtained from the peripheral blood mononuclear cells (PBMCs) of a significant group of Italian intravenous drug users (IVDUs), prison inmates and blood donors (BDs) who were HTLV antibody positive. Restriction fragment length polymorphism (RFLP) of amplified LTR region with ApaI, NdeI, DraI, SacI and MaeIII endonucleases was used to define the HTLV-I subtypes, while the different variants of HTLV-IIa and -IIb were defined by RFLP of the LTR region with the AvaII, BglI, SauI, XhoI and BanII endonucleases. The four HTLV-I isolated from BDs were characterized as C type. All the 11 HTLV-II detected in the IVDUs were HTLV-IIb4, while among the prisoners one HTLV-IIb5 and five HTLV-IIb4 were found. Interestingly, in the BDs group two HTLV-IIa0 and one HTLV-IIb4 were detected. It should also be noted that 82% of the IVDUs and 50% of the prisoners were coinfected with HIV, while all the BDs were HIV negative. These data indicate that HTLV-IIb4 is the predominant genotype in Italian IVDUs and prisoners, while the significant variability observed in the BD HTLV-II isolates could be due to the different source of infection among this group.

Nucleotide sequence and restriction fragment-length polymorphism analysis of human T-cell lymphotropic virus type II (HTLV-II) in southern Europe: evidence for the HTLV-IIa and HTLV-IIb subtypes.

Human T-cell lymphotropic virus type II (HTLV-II) has been subtyped into two major groups, IIa and IIb, according to molecular studies involving env gene sequencing. Subsequently, this retrovirus was further subclassified by examining the long terminal repeat (LTR), the most divergent genomic region. Sequence analysis and restriction fragment-length polymorphism (RFLP) applied to the LTR region identified either four or five groups within the IIa subtype (depending on the restriction enzyme sets used) and six within the IIb subtype. In this study, we analyzed the LTR sequences of 29 samples obtained from HTLV-II-infected individuals living in Spain and Italy, which included 24 injecting drug users (IDUs), three blood donors, and two subjects at risk for HIV/HTLV infection. Sequence analysis and phylogenetic analysis of 720 base pairs of the LTR performed in 10 Spanish samples showed that all of these samples belonged to IIb subtype, with a divergence of 7.5% and 1.66% compared with MoT (IIa) and NRA/G12 (IIb) isolates, respectively. RFLP analysis demonstrated the presence of the IIb 4-subtype restriction pattern in 26 samples, a IIb5-subtype pattern in one Italian IDU, and a IIa0-subtype pattern in two Italian samples (blood donors), according to W.M. Switzer's nomenclature. This is the first report of the presence of IIb5 in Southern Europe and IIa0 among Italian blood donors. RFLP correlated with nucleotide sequence and phylogenetic data obtained in this study, demonstrating the ability of the RFLP method to predict the phylogroup of HTLV-II-infected samples.

A multicentric seroepidemiological survey of HTLV-I/II in Italy.

BACKGROUND: Several studies carried out in the USA and in Europe have shown the presence of HTLV-I/II antibodies in subjects belonging to high-risk groups for HIV infection as well as blood donors. Concern about the presence of HTLV-I/II markers in the normal population, as well as the efficient transmission of HTLV-I/II by whole blood or infected blood cells have led several countries to include screening for anti-HTLV-I/II among the mandatory serological testing of blood donors. OBJECTIVE: In order to assess the risk of HTLV-I/II infection related to blood transfusions, a multicentric survey for antibodies against HTLV-I and HTLV-II was carried out involving 10 Italian sites during the spring of 1991. STUDY DESIGN: Serum specimens were collected from 14,598 blood donors, 1,411 injecting drug users, 1,015 thalassemics, 142 hemophiliacs and 138 hemodialysis patients. HTLV antibodies were detected by a screening EIA which combines a viral lysate with a recombinant HTLV-I env protein (p21e). The serological confirmation was performed by a semi-automated dot-blot immunoassay that detects gag p19 and p24 and env p21e specific antibodies, while the discrimination of HTLV-I and HTLV-II reactivities was carried out by EIAs employing synthetic peptides of the ENV region specific for each virus. RESULTS: The seroprevalence of confirmed positives was 0.034% among blood donors and 3.61% among IDUs, while no sample of the other categories could be confirmed, although several were indeterminate and one thalassemic reacted against HTLV-I on peptide testing. HTLV-I reactivity was observed in one blood donor, while all 38 of the 51 confirmed seropositive IDU's reacted only to the HTLV-II synthetic peptide. CONCLUSIONS: These data confirm a high prevalence of HTLV-II among Italian IDUs and show an HTLV-I/II seroprevalence among blood donors very similar to that which was found in the USA volunteer blood donors. A surveillance program among blood donors seems advisable in order to establish the possible need of a mandatory screening for HTLV-I/II.

Bolus vs. continuous hepatic arterial infusion of cisplatin plus intravenous 5-fluorouracil chemotherapy for unresectable colorectal metastases.

UNLABELLED: A multicenter, randomized Phase 2 study that compared patients, affected by colorectal liver metastases, who received intrahepatic arterial infusion with two different schedules of cisplatin, bolus vs. continuous infusion, and systemic 5-fluorouracil. PURPOSE: The aim of this study was to validate results of a previous Phase 2 trial on bolus cisplatin intrahepatic arterial infusion, which reported a 47 percent response rate and a 32 percent 4-year survival rate for Gennari's Stage 2 patients, with a high rate of neurologic, gastrointestinal, and hematologic toxicity. METHODS: One hundred nine patients were randomized in a Phase 2 study to receive cisplatin intrahepatic arterial infusion (24 mg/m2/day, 1-->5, bolus vs. continuous infusion) and systemic intravenous 5-fluorouracil (250, 375, or 500 mg/m2/day, 1-->5; escalating doses, respectively, at cycles I, II, III, and VI). To avoid neurotoxicity a maximum of six cycles was administered. RESULTS: Preliminary results for the 78 evaluable patients are similar to those of the previous study: response rate 46 percent and at a median follow-up of 16.5 months, the overall survival was 16.5 months, with 45 percent of the patients who received more than 3 cycles alive at 3 years. Toxicity, evaluable in 99 patients, showed a decreased incidence of neurotoxicity and a tolerable gastrointestinal and hematologic toxicity, lower in the cisplatin continuous infusion arm. CONCLUSION: This study clearly shows that cisplatin intrahepatic arterial infusion is able to provide a good palliative effect with a tolerable toxicity.

Nerve-sparing surgery in 302 resectable rectosigmoid cancer patients: genitourinary morbidity and 10-year survival.

PURPOSE: The aim of this study was to evaluate 5-year and 10-year disease-free survival, urinary dysfunction, and sexual activity after nerve-sparing radical surgery, including lumboaortic lymphadenectomy for rectosigmoid cancer. METHODS: Since 1980 to 1992, 302 consecutive patients affected with rectal (188) or sigmoid (114) resectable cancer underwent radical surgery. Lumboaortic lymphadenectomy was routinely performed and total mesorectal dissection was always accomplished in rectal cancer. Splanchnic nerves, superior hypogastric plexus, hypogastric nerves, and sacral parasympathetic nerves were sought, identified, and preserved or, when necessary, unilaterally sacrificed. Fifty-three (17.6 percent) patients were classified Dukes A, 145 (48.0 percent) B, 46 (15.2 percent) C1, and 17 (5.6 percent) C2. Thirty-nine (12.9 percent) patients were Dukes D. In 85 rectal cancer patients, tumor was located at the lower third. Eighty-six of 210 Dukes B and C patients were submitted to systemic chemotherapy and/or high-dose pelvic radiotherapy. RESULTS: The actuarial 5-year disease-free survival was 58.5 percent in rectal and 65.7 percent in sigmoid cancer patients, median follow-up time was 47 months. During the follow-up, each patient was interviewed about sexual activity and urinary dysfunction and a questionnaire was filled out. Urinary dysfunction was not frequently observed, while a definitive sexual impotence was reported in 27.6 percent of the patients. The age under 60 years and sphincter-saving surgery were demonstrated as significantly contributing to retaining a satisfactory sexual activity. CONCLUSIONS: Unexpectedly high disease-free survival was observed in the Dukes C2 subgroup. It allows us to hypothesize that lumboaortic lymphadenectomy could remove neoplastic microfoci present at this level in those patients, enhancing surgical chances of cure. The majority of male patients under 60 years old can retain a satisfactory sexual activity after undergoing a nerve-sparing sphincter-saving cancer surgery.

[The use of an ultrasonic lithotripter during video laparoscopic cholecystectomy]. / Uso del litotritore ad ultrasuoni in corso di colecistectomia videolaparoscopica.

The video laparoscopic cholecystectomy, a new technique recently introduced in surgical surgery, includes, among other complications, also the dimension of stones. Our intention in the present work is to remove this limitation using an ultrasound lithotripter to reduce the dimension of lithiasic formations, avoiding, after all, to resort to minilaparotomy or to the use of dilators which are in contrast at least with two of the principles of methodology, the aesthetic and functional one for the patient.

Genetic alteration in gastrointestinal cancer. A molecular and cytogenetic study.

We examined 16 cases of gastrointestinal cancer, of which 11 were from the colon, 1 from the rectum, and 4 of gastric origin, cytogenetically for expression and for loss of heterozygosity (LOH) on chromosome 18 using Deleted Colon Cancer (DCC) gene. LOH on chromosome 18 with DCC probe was detected in 7 out of 11 cases of colon, in 4 out of 4 cases of gastric and in 1 case of rectum cancer. In all gastrointestinal tumors the expression of DCC gene was absent, while it was present in normal tissue. We also found rearrangements of chromosomes 18 (10 cases) and 17 (9 cases), leading respectively to deletions of long and short arms. Other additional abnormalities were observed involving chromosomes 5, 6, 15 and 19. The data recorded in our series differ from other authors' data in three respects: a high incidence of pseudodiploid chromosome number, rearrangements of chromosome 19 and 15, and involvement of DCC gene in the development of gastric cancer, as well as in colorectal cancer as previously reported.

TAG-72 (CA 72-4 assay) as a complementary serum tumor antigen to carcinoembryonic antigen in monitoring patients with colorectal cancer.

BACKGROUND: Serum carcinoembryonic antigen (CEA) is the most frequently chosen tumor marker in the clinical diagnosis of colorectal carcinoma and in the long-term monitoring of patients after tumor resection. In recent years, monoclonal antibody technology has identified several new markers of neoplasia, two of which, TAG-72 and CA 19-9, are found in the sera of patients with adenocarcinoma. Serum CEA, TAG-72, and CA 19-9 were evaluated in 300 patients with either malignant (n = 200) or benign (n = 100) colorectal disease. METHODS: Serum CEA, TAG-72 (CA 72-4), and CA 19-9 antigen levels were determined with a double-determinant radioimmunometric assay kit. Samples and appropriate standards were assayed in duplicate. The cutoff limits used for each assay were indicated by the manufacturer. All of the results of the CA 72-4, CEA, and CA 19-9 serum assays were separated from the clinical information until the study was completed. RESULTS: Of the 200 patients with colorectal carcinoma, the percentage of patients whose serum samples were positive for CEA, TAG-72, or CA 19-9 was 43%, 43%, and 27%, respectively. The measurement of TAG-72 with CEA for patients with primary or recurrent colorectal carcinoma increased substantially (to 60%) the percentage of positive serum samples when compared with measuring each serum tumor marker alone. Moreover, the apparent advantage gained by measuring the two tumor markers was achieved with little increase in the number of false-positive results. CONCLUSIONS: The findings support previous observations of complementary expression of TAG-72 and CEA and indicate that a significant advantage could be gained in the detection of primary and, perhaps, recurrent colorectal carcinoma by incorporating the measurement of serum TAG-72 with that of CEA.

The role of the administration time of prophylactic antibiotic therapy in colorectal cancer surgery: a review of 6,069 patients from 36 randomized clinical trials.

The aim of the present report was to establish the effectiveness of different prophylactic antibiotic regimens and administration times in colorectal cancer surgery. Six thousand and sixty nine patients from 36 selected randomized clinical trials, published between 1980 and 1989, were reviewed. The occurrence of septic events, isolated bacterial strains, fever and postoperative hospitalization times were also analyzed. The therapeutic schedules that included the perioperative administration of antibiotics provided better results that those that did not (p. less than .0001 for infections both specifically related and unrelated to colorectal surgery). The number of postoperative administrations did not affect the clinical results, even if the predominant choice was to give more than one administration of antibiotics. A factorial design demonstrated that prolonging the perioperative administrations up to the postoperative period provided statistically significant benefits (p less than .0001) only with regard to the risk of infections that were not specifically related to colorectal surgery.

Flow cytometric analysis of DNA content and cell kinetics in colorectal carcinoma.

DNA ploidy and cell proliferation were studied by means of flow cytometry in 98 patients with primary colorectal adenocarcinoma. Multiple samples of tumour burden were pooled and freshly dissociated immediately after surgery for FACS analysis. The relationships between ploidy, proliferative activity, evaluated in terms of S-phase percentages (%S), and some clinico-pathological variables were analyzed. 87 of the 98 tumors yielded evaluable DNA histograms: 32 were diploid (37%) and 55 were aneuploid (63%; median DNA index = 1.6). Multiple aneuploid cell populations were found in 15 tumors (17%). The % S was estimated by means of a mathematical model. Aneuploid tumors showed % S values significantly higher than diploid ones (p < 0.0001). Differences in the distribution of DNA aneuploidy were observed in relation to Dukes' stage and tumor site, left colon, rectum and stage D tumors being more frequently aneuploid. No significant differences in proliferative activity were observed in relation to most of the clinical variables, except for higher % S values observed in tumors of right colon compared to those of left colon and rectum.

[Chronic exposure to inhalation anesthetics and immunity]. / Esposizione cronica agli anestetici inalatori ed immunità.

The existence of a professional disease caused by exposure to general anesthetics has been reported by a number of studies, but opinions still differ as to the effective implications of this type of exposure. The aim of the present study was to analyse alterations in cellular and humoral immunity in anesthetists regularly exposed to general anesthetics. Regression analysis showed that IgA and NK lymphocytes were significantly increased whereas total T lymphocytes reduced in parallel with age and length of service. It can therefore be supposed that the increase in IgA might suggest chronic liver disease, while alterations in the lymphocyte populations, even if closely associated to chronic exposure leading to the hypothesis of a cause-and-effect relationship, do not alter immune system functions since they are similar to those that occur with ageing.

A tetrazolium-based colorimetric assay for quantification of HIV-1-induced cytopathogenicity in monocyte-macrophages exposed to macrophage-colony-stimulating factor.

A sensitive assay was developed for in vitro evaluation of anti-HIV agents in monocyte-macrophage cells (M/M) (a crucial target of HIV in the body). Monocyte-macrophage cells are usually poorly sensitive to the cytopathic effect induced by HIV. However, when fresh adherent monocyte-macrophage cells are cultured at relatively high density in the presence of macrophage-colony stimulating factor (M-CSF), they undergo cytolysis and die in 2-3 weeks. HIV-mediated cell-killing can thus be assessed with a method based on the reduction of the yellow colored 3-(4-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) by metabolically active cells to a blue formazan, which can be measured spectrophotometrically. HIV-mediated cytopathic effect of M-CSF-exposed monocyte-macrophage cells was consistently achieved in all experiments performed under the conditions described herein. Anti-HIV activity of zidovudine (AZT) was also comparatively evaluated in M-CSF- and normal monocyte-macrophage cells both using the MTT assay and by measuring HIV-p24 antigen production in supernatants of monocyte-macrophage cells cultures, and similar results obtained with both methods. These results support the use of this colorimetric assay for broad screening of anti-HIV agents in monocyte-macrophage cells.

Tumor-associated glycoprotein-72 serum levels complement carcinoembryonic antigen levels in monitoring patients with gastrointestinal carcinoma. A longitudinal study.

Eighty-two patients diagnosed with gastrointestinal (GI) adenocarcinoma were evaluated before and for 26 months after primary tumor resection for the presence of two serum tumor markers: tumor-associated glycoprotein-72 (TAG-72) and carcinoembryonic antigen (CEA). Elevated TAG-72 and CEA serum levels were found preoperatively in 32 (39%) and 34 (41.5%) of the 82 patients, respectively. The percentage of patients with elevated serum levels of either TAG-72 or CEA was 56.1% (46 of 82). Twelve (15%) patients who had normal CEA serum levels had elevated TAG-72 serum levels, and conversely, serum from 14 (17%) patients who were TAG-72 negative were CEA positive. Forty-five of the 82 patients were diagnosed with advanced disease (i.e., Stages C and D for colorectal, Stages III and IV for stomach), and 29 (64.4%) and 26 (57.8%) of those patients had elevated serum levels of TAG-72 or CEA, respectively. Elevated levels of either TAG-72 or CEA, however, were found in sera of 82.2% of patients with advanced GI cancer, which is an increase of 24.4% over the use of CEA antigen alone as a marker of disease. The measurement of both TAG-72 and CEA may improve the diagnosis of patients with GI malignant disease due to the apparent complementary association which exists between these tumor markers. Serum TAG-72 and CEA levels were monitored in 31 patients for varying lengths of time after resection of the carcinoma; 11 patients developed recurrent disease. Sera from nine of 11 (81.8%) of these patients had elevated TAG-72 levels and six of 11 (54.5%) had elevated CEA levels. Tumor marker elevations were observed either before (35 to 166 days) or at the time of diagnosis of recurrence. The elevation of one or both markers correlated with the clinical status in ten of 11 (90.9%) patients with recurrence. In addition, 20 patients who were clinically free of disease after more than 700 days' follow-up had normal serum levels of both TAG-72 and CEA. These findings suggest that the combined use of serum TAG-72 and CEA measurements may improve detection of recurrence in patients with GI cancer and may be useful in the postsurgical management of GI adenocarcinoma patients.

Persistence of HIV-1 silent infection in seronegative subjects at high risk.

Twenty regular sexual partners of HIV-1 infected subjects, without detectable human immunodeficiency virus (HIV-1) antibody and positive for HIV-1 genome by in situ hybridization (ISH), were selected and studied longitudinally for 6-36 months to estimate the duration of silent infection. During the follow-up period, 10 showed atypical Western Blot (WB) patterns. Two seronegative partners seroconverted. Rapid progress to AIDS was observed in 7 seropositive subjects.