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Background The role of Mycobacterium avium paratuberculosis [MAP] in inflammatory bowel disease [IBD], especially Crohn’s disease [CD] is controversial due conflicting results and lack of reproducibility and standardised tests. The current study focuses on the role of MAP in disease progression and genetic susceptibility, as MAP is likely one of many factors involved in the complex pathogenesis of IBD, potentially affecting a subgroup depending on genetic susceptibility. Methods Serum from 812 patients was evaluated with seven immunoglobulin [Ig] isotype-specific serology tests assessing humoral response to three different MAP antigens. For each of these in total 21 tests, the intra-assay and inter-assay coefficients were used to evaluate test accuracy. Reliable assays were subsequently analysed in relation to disease characteristics and need for biologic therapy/surgery. Genome-wide genotyping was available for all participants. Genetic determinants of humoral response to MAP antigens were evaluated using genome-wide association analysis and polygenic risk scores [PRS]. Results High IgA or IgM response to MAP2609 was associated with increased use of biologic therapy in CD and ulcerative colitis [UC] [odds ratios 2.69; 95% confidence interval 1.44–5.01; and 2.60, 1.46–4.64, respectively]. No associations were seen for risk of surgery [p-values > 0.29]. We could not identify genetic determinants nor polygenic risk scores for MAP response with genome-wide significance. Conclusions Extensive assays for serological response to MAP were evaluated using stringent criteria for reliability. Increased IgA and IgM response to MAP antigens was seen in patients exposed to biologic therapy, but no genetic determinants underlying this humoral response were found.

Background Ustekinumab (UST), which targets p40 of IL23/IL12, is an effective treatment for Crohn’s disease (CD). Therapeutic drug monitoring (TDM) may optimize UST posology. The aim of this study was to investigate UST and IL23 serum and tissue concentrations in relation to mucosal inflammation and treatment response. Methods A prospective cohort study was performed in adult CD patients who initiated UST at Erasmus MC, the Netherlands, between December 2016 and November 2018. Endoscopies were performed at baseline and week 16. UST and IL23 serum and tissue concentrations were measured at week 16. Clinical and biochemical response were defined as decline of ≥3 points in Harvey Bradshaw Index and reduction of ≥50% in fecal calprotectin levels. Endoscopic response was defined as a ≥50% decline in simple endoscopic score or a decline of ≥1 points in Rutgeerts’ score. Histological remission was defined as GHAS score ≤4. Results All 56 included patients were previously treated with anti-TNFα-inhibitors prior to UST. Upon treatment with UST, 17 (30%) showed clinical response, 16/53 (30%) biochemical response, and 20/56 (36%) endoscopic response. UST, but not IL23, concentration in biopsies was correlated to levels in corresponding sera (p < 0.0001). No correlation was found between UST serum and tissue levels with clinical or endoscopic response, or histological remission. However, patients achieving biochemical response showed significantly higher UST serum levels as compared to those without biochemical response (3.12 µg/ml vs 1.41 µg/ml, p = 0.01). Furthermore, tissue IL23/UST ratio correlated with histologic mucosal inflammation (p = 0.01). Conclusion This is the first study to demonstrate a correlation between serum and tissue UST levels. We showed that the tissue IL23/UST ratio correlated with mucosal inflammation. UST tissue concentrations did not correlate with treatment response, while UST serum concentrations were associated with biochemical response at week 16. The role of UST levels for TDM in IBD warrants further research.

Background: This study compared the effects of insulin pump (IP) vs. multiple daily injection therapy (MDI) on glucose variability (GV) evaluated by professional Continuous Glucose Monitoring (CGM) in real-life condition of patients with type 1 diabetes (T1D). Methods: This was a parallel study in 120 T1D patients, with a mean age of 31.2 years, and hemoglobin A1c (HbA1c) level of 8.3%; 99 patients were treated MDI (66 patients with Glargine U100, 13 with Detemir, 20 with Degludec U100) and 21 with IP. The primary end point was the coefficient of variation (CV) of blood glucose. Secondary end points included HbA1c, mean amplitude of glycemic excursions (MAGE). We compared IP vs. MDI. Results: IP was not different to MDI respect to CV (37.94±1.68 vs. 40.32±075; p=0.12) and HbA1c (8.59±0.20 vs. 8.23±0.15; p=0.39). There were also no significant differences between treatment groups with respect to MAGE (308.75±11.28 vs. 309.22±5.17; p=0.92) and total doses of insulin (35.14±4.77 vs. 42.88±2,22; p=0.36). In multiple regression analyses, the CV was positively associated with MAGE and negatively with HbA1c only 22% (R2: 0,22, p = 0.01). Conclusion: In our population in real world data, no significant difference was observed in the GV indicators in patients treated with MDI vs. IP. Disclosure A.E. Proietti: None. A.V. Daghero: None. M.L. Scapuzzi: None. M.L. Iglesias: None. J.P. Nogueira: None.

Background: This study compared the effects of insulin glargine and insulin detemir or degludec on glucose variability with CGM technology in real-life condition of patients with T1D. Methods: This was a paralell trial in 81 T1D patients: 27 men and 51 women, with an average age of 31.33 years, and hemoglobin A1c (HbA1c) level of 8.4%. 56 patients were treated with glargine and 25 with detemir/degludec. The primary end point was the coefficient of variation (CV) of blood glucose. Secondary end points included HbA1c, mean amplitude of glycemic excursions (MAGE) and total doses of basal insulin and rapid analogs. Results: insulin degludec/detemir was not different to insulin glargine with respect to CV (39.42±7.64 vs. 40.37±7.38, p=0.95) and HbA1c (8.03±1.42 vs. 8.47±1.68, p=0.26). There were also no significant differences between treatment groups with respect to MAGE (305.77±7.50 vs. 300.83±10.64, p=0.71) and rapids analogs (17.92±1.34 vs. 17.50±1.88, p=0.85). Only significantly higher doses were found with insulin detemir/degludec vs. insulin glargine (33.30±5.60 vs. 21.70±1.53, p=0.01). In multiple regression analyses, the total dose of basal insulin was the only parameter positively associated with levels of HbA1c (p = 0.01). Conclusion: the increased doses of insulin determir/degludec related to insulin glargine do not lead to a better metabolic control regarding CV or HbA1c levels. Disclosure A.E. Proietti: None. A.V. Daghero: None. N. Panei: None. M.L. Scapuzzi: None. A.E. Jokiel: None. M. Iglesias: None. J. Velazquez: None. J.P. Nogueira: None.

The interaction of two different phenomena, the gas diffusion in a membrane and the spatial dependent sensitivity of a transducer, affects the final response of a sensor. In this paper we present data and simulation of a quartz crystal microbalance (QCM) utilizing a thin film of palladium as hydrogen detector. Preliminary results show that it is possible to exploit these peculiar properties of the membrane and of the transducer to enhance the overall performance of the sensor. The membrane and transducer used for this work are merely one example and the concept here exposed can be applied to other more complex systems.

This paper deals with problems of both molecular compounds and fine parti cles present as pollutants in surrounding environment. Criteria classification as defined by Italian law are presented in terms of exposure limit as the relc:vant parameter. Also a list of detection methods and detection limits is presented. This reference may help in the sensor strategy assessment for environmentill eonu-ot Finally a block diagram indicating the environmental evolution due to the possible energy transformation is bere presented and discussed in order to emphasise that pollutants do represent by-products of industriai processes. Our duty is to reduce as much as possible the concentration of these pollutants. Whatever the solution taken, sensors will always play a fundamental role for monitoring purposes.

This study investigates the hypothesis that the nonlinear component of human heart rate (HR) variability might show a periodic structure over the 24-h span. Such a postulate could explain how the chaotic component might coexist with the deterministic periodic variability of instantaneous HR in beat per minute.The sinusal R-R intervals (sRRi) of the Holter EKG of 10 clinically healthy subjects (5 M, 5 F, 23-30 years) were analyzed per each hour of the day-night span according to two methods for the nonlinear chaotic variability, i.e., the correlation dimension method, and the linear periodic variability, i.e., periodic regression analysis.The hourly-qualified correlation integrals were found to show a significant circadian rhythm, with an acrophase located during the night in coincidence with the longest duration of the sRRi and the lowest rate of cardiac pulse.The rhythmic structure of the chaotic component of the human HR variability let us to think that a deterministic periodic chaos of fractal type regulates the nonlinear cardiac dynamics. Such a periodic structure allows the chaos to be compatible with the deterministic linear periodicity of circadian type which characterizes the within-day variability of human HR.

The residual resistance of severe combined immunodeficiency (SCID) mice to human graft is the main factor in conditioning both the extent of human cell reconstitution and the xenograft-to-xenograft variability. We have recently shown that an early and massive murine granulocyte recruitment is the main event in the SCID mouse reaction to the human graft.Here, we evaluate the importance of mouse granulocytes in the restriction of human cell engraftment in SCID mice. We injected SCID mice with a monoclonal antibody to murine granulocytes.Injection of this antibody resulted in a marked depletion of polymorphonuclear cells in the hematopoietic organs of SCID mice. This depletion was associated with a significant increase in both the growth of human cell lines of different hematopoietic origin and the engraftment of human peripheral blood leukocytes. Moreover, the abolishment of the early granulocyte reaction markedly reduced the xenograft-to-xenograft variation, a major shortcoming of these xenochimeric models.These results provide new insights into the control of the natural immune response of SCID mice against human graft. Furthermore, treatments aimed at controlling the acute inflammatory reaction of SCID mouse-to-human cell transplantation can be considered useful experimental approaches for increasing the xenograft-to-xenograft reproducibility.

The aim of this study was to investigate the circadian variability of heart rate in acute myocardial infarction (AMI) in identifying patients at high risk for malignant ventricular arrhythmias (MVA) and sudden death within 1 year of the acute event. The investigation was carried out in 43 patients, who underwent 24-hour Holter monitoring within 3 months of AMI. Besides the time domain indexes of heart rate variability (SDNN, SDNN index, pNN50, rMSSD), the circadian rhythm of hourly total beats (HTB) and hourly qualified beats (HQB) has been analyzed by the Cosinor method. The AMI patients with MVA and those with MVA who died within 1 year the acute event showed SDNN, SDNN index and pNN50 values lower than subjects without MVA and survived patients with MVA, respectively; the individuals with AMI at high risk for MVA and for sudden death had an SDNN value105 ms and 50 ms, respectively. The circadian rhythm of HTB and HQB was statistically validated only in the group without MVA; patients without the circadian rhythm of HTB and HQB showed a higher mortality rate within 1 year of AMI, and the majority was in the group with MVA. The contemporary evidence of an SDNN value105 ms and the lack of HTB and HQB circadian rhythm increased sensitivity for identifying patients with MVA to 75%. On the other hand, the contemporary evidence of an SDNN value50 ms and the lack of HTB and HQB circadian rhythm increased sensitivity for identifying patients who died within 1 year of AMI to 100%. In conclusion, the assayed methods seem to be both useful and complementary in identifying patients at high risk for MVA and sudden death within 1 year of AMI.

The TAL-1 gene specifies for a basic domain-helix-loop-helix protein, which is involved in the control of normal hematopoiesis. In human pathology, the TAL-1 gene product is expressed in a high percentage of T-cell acute lymphoblastic leukemias in the pediatric age range; however, it has not been established whether the expression has a causal role in oncogenesis. In this report, we describe the phenotype of mouse transgenic lines obtained by inducing tal-1 protein expression in lymphoid tissues using the LCK promoter. The survival rate of tal-1 transgenic animals was much lower as compared with control mice. Histopathological analysis revealed lymphomas of T-cell type, often comprising a minor B-cell component. Some mice showed marked splenic lymphocyte depletion. Primary lymphocyte cultures showed partial independence from exogenous growth stimuli and increased resistance to low-serum apoptosis. To further unravel the tal-1 oncogenic potential, a strain of tal-1 transgenic mice was crossbred with p53-/- mice; the survival rate in these animals was reduced by more than one-half when compared with that of tal-1 mice, and histopathological analysis revealed exclusively T-cell lymphomas. These data indicate that TAL-1, expressed in T cells, is per se a potent oncogene, which may exert a key leukemogenetic role in the majority of T-cell acute lymphoblastic leukemias.

We have studied the effects of single and combined treatment with recombinant human interleukin 1 beta (IL-1 beta) and recombinant human interleukin-6 (IL-6) on spleen and bone marrow hematopoiesis in normal and cyclophosphamide-treated mice. Injection of IL-1 beta alone resulted in a significant increase in the number of granulocytes and splenic progenitors [burst-forming units-erythroid (BFU-E) and colony-forming units-granulomonocytic (CFU-GM)] as compared with control mice but did not markedly enhance the number of bone marrow BFU-E and CFU-GM. IL-6 alone had little effect on the number of splenic progenitors but significantly increased the number of marrow BFU-E and CFU-GM, especially after a 6-day cytokine treatment. Combined daily administration of IL-1 beta and IL-6 for 3 days resulted in a synergistic stimulation of hematopoiesis as evaluated by the number of spleen and bone marrow CFU-GM and BFU-E colonies. Likewise, IL-1 beta/IL-6 markedly enhanced the number of circulating neutrophils, whereas each cytokine alone had little or no effect. When the numbers of spleen progenitors and peripheral granulocytes were determined 1 day after the last injection, a synergistic myelostimulatory effect of combined IL-1 beta/IL-6 treatment was observed at all doses (IL-1 beta, 0.25-0.5 microgram; IL-6, 1-20 micrograms). Furthermore, combined treatment with IL-1 beta/IL-6 accelerated and potentiated the recovery of myeloid cells after cyclophosphamide injection, whereas the single regimen treatment was not effective. Particularly, the rebound of WBC (especially neutrophilic granulocytes) after cyclophosphamide treatment was markedly enhanced by the combined treatment, whereas the single regimen was ineffective. Altogether these results may contribute to the development of combination therapies with cytokines and antiblastic agents in the treatment of cancer patients.

Highly metastatic alpha/beta-interferon (IFN-alpha/beta)-resistant Friend leukemia cells (FLC) were transfected with a retroviral vector (pLTneoL-5) containing the mouse IFN-alpha 1 gene. Transfected clones were isolated and tested for their capacity to secrete IFN-alpha 1 and their tumorigenicity when injected s.c. into immunocompetent syngeneic DBA/2 mice. Almost all FLC clones producing IFN in the range of 16-512 units/ml failed to grow when injected s.c. or i.p. into normal mice, whereas control FLC (transfected with a vector without the IFN gene) exhibited the highly malignant phenotype of the original FLC. High levels of IFN were detected in peritoneal fluid, tumor extracts, and sera of mice given injections of IFN-producing cells. Injection of mice with antibodies to IFN-alpha/beta resulted in the development of tumor ascites in mice transplanted i.p. with IFN-producing FLC. In contrast to the tumor rejection observed in immunocompetent mice, IFN-producing FLC were highly tumorigenic when transplanted into immunosuppressed nude mice. Mice given injections of IFN-producing FLC developed a long-lasting tumor-specific immune resistance to subsequent injection with highly metastatic FLC. Simultaneous s.c. injection of both metastatic FLC (approximately 10(3) 50% lethal doses) and IFN-producing cells resulted in potent inhibition of the tumor growth, with a survival rate of approximately 50% for injected mice. Contralateral injection (s.c.) of IFN-producing FLC into mice with established metastatic tumors produced a marked inhibition of tumor growth, with a survival rate of 10% for injected mice. These results indicate that: (a) the genetic modification of highly metastatic FLC by means of transfer of the IFN-alpha 1 gene results in potent tumor cell rejection, which is mediated by an IFN-induced host immune response; (b) injections of IFN-producing tumor cells are effective in inhibiting tumor growth in mice with established metastatic tumors. These data suggest that tumor cells transfected with the IFN-alpha gene might be used as an effective therapy for the treatment of certain human metastatic tumors, provided that suitable strategies are defined to prevent growth of the cytokine-producing cells.

We have studied the effects of single and combined treatment with interleukin 1 beta (IL-1 beta) and interleukin 2 (IL-2) on spleen and bone marrow hematopoiesis in normal mice. Injection of IL-1 beta alone was followed by a significant increase in the number of granulocytes in spleen and progenitors (burst-forming units-erythroid and colony-forming units-granulomonocytic) in both spleen and bone marrow, s compared to control mice. In contrast, IL-2 alone induced only a slight increase in the number of marrow colony-forming units-granulomonocytic and had no significant effect on spleen progenitors. Repeated injections of both IL-1 beta and IL-2 resulted in a synergistic increase in spleen weight and splenocyte number, as compared to mice treated with the single cytokine regimen; in particular, the combined treatment induced a marked rise in the number of neutrophilic granulocytes and erythroblasts, whereas splenic lymphocytes were not affected. This regimen also caused a synergistic increase in the number of spleen and marrow progenitor cells: a time-course analysis showed an elevation in numbers of both burst-forming units-erythroid and colony-forming units-granulomonocytic, first in marrow (day 10) and subsequently in spleen (day 18). Combined IL-1 beta/IL-2 treatment dampened the decrease and accelerated the recovery of myeloid cells after cyclophosphamide injection, whereas the single cytokine regimen was not effective. Similarly, the rebound of WBC (especially neutrophilic granulocytes) after cyclophosphamide treatment was markedly enhanced by the combined treatment, whereas the single cytokine regimen was ineffective. These results, indicating a myelostimulatory effect by the combined cytokine regimen, together with our previous observations showing a synergistic antitumor activity by IL-1/IL-2 treatment in experimental mouse tumors (V. Ciolli et al., J. Exp. Med., 173: 313-322, 1991), may provide the basis for the development of new combination therapies with cytokines and antiblastic agents in the treatment of cancer patients.

In this study, we describe the origin and characterization of a new metastatic tumor cell line (p11-R-Eb) obtained after i.p. passages of the nonmetastatic Eb lymphoma cells into DBA/2 mice. The p11-R-Eb cells exhibited the same morphology and in vitro growth properties and chromosome markers as the original Eb cells. FACS analysis of the p11-R-Eb cells also revealed a close similarity to the Eb cells. Moreover, the p11-R-Eb cells were specifically killed by anti-Eb cytotoxic lymphocytes. In spite of all these characteristics of the Eb line, p11-R-Eb cells metastasized to the liver when injected i.v. or s.c. in DBA/2 mice. Peritumoral interleukin (IL)-2 treatment resulted in a potent antitumor response in DBA/2 mice transplanted s.c. with p11-R-Eb cells. In contrast, the same IL-2 regimen did not significantly increase the survival time of mice transplanted with the highly metastatic ESb cell line. Combined IL-1/IL-2 treatments of established p11-R-Eb tumors resulted in a synergistic antitumor effect and in tumor regression in 70% of the injected mice. Similarly, combined peritumoral treatment with IL-1 and interferon-alpha/beta, which were poorly effective or ineffective as single cytokine therapy, resulted in a marked antitumor effect, and 30% of the mice were cured. Spleen cells from IL-1/IL-2-treated p11-R-Eb-cell-injected mice showed a marked antitumor activity when assayed in a Winn assay with homologous tumor cells. This antitumor activity was eliminated by preincubation of spleen cells with antibodies to CD4 and complement and markedly inhibited by anti-asialo GM1 antibodies. P11-R-Eb cells represent, therefore, a new tumor model which may be useful for investigating the relevant mechanisms which need to be activated to achieve a potent antitumor response to cytokine therapy in the DBA/2 mouse host.

Administration of alpha/beta-interferon (IFN) exerts a marked antitumor effect in DBA/2 mice given injections i.v. of large numbers of IFN-alpha/beta-resistant erythroleukemia cells (FLC). To investigate the possible mechanisms of FLC tumor inhibition in the liver of interferon-treated mice, we developed an in vitro model consisting of a coculture of IFN-alpha/beta-resistant 3Cl8 FLC and syngeneic mouse hepatocytes. Whereas IFN-alpha/beta did not inhibit the multiplication of 3Cl8 FLC cultivated alone, it effectively inhibited the multiplication of 3Cl8 FLC in coculture with hepatocytes. The inhibitory effect was directly proportional to the amount of IFN-alpha/beta added to the cocultures, and more than 90% inhibition of FLC multiplication was noted with 1.6 x 10(5) IU/ml of IFN-alpha/beta on Day 3 of coculture. When FLC were separated from the monolayer of hepatocytes by a pored membrane (0.4 microns), the inhibitory effect on FLC proliferation was unchanged, indicating that a soluble factor(s) released from IFN-treated hepatocytes was most important in the inhibition of FLC multiplication. An inhibitory activity of FLC multiplication was detected only in the conditioned medium of IFN-treated hepatocytes but not in the conditioned medium of control hepatocytes nor in extracts of IFN-treated or control hepatocytes. The inhibitory factor(s) in the conditioned medium of IFN-treated hepatocytes was retained by an ultrafiltration membrane (Mr cut off 10,000), and its activity was completely abrogated by trypsin digestion. Its stability to treatment with 1 M acetic acid as well as lack of correlation between the antiproliferative effect and the amount of L-arginine in the medium distinguished this factor(s) from liver arginase which was also found to be a potent inhibitor of FLC multiplication in vitro. The inhibitory factor(s) was also distinguishable in its biological activity from IFN gamma, interleukin 1 alpha and beta, and transforming growth factor beta 1 and beta 2. These results suggest the possibility that the inhibitory effect of IFN-alpha/beta on the development of 3Cl8 FLC in the livers of IFN-treated mice may be mediated by an IFN-induced inhibitor of FLC multiplication.

1080 Background: X has demonstrated consistently high single-agent activity and good tolerability in pretreated and chemonaive MBC, and extends survival when added to docetaxel. High activity, minimal myelosuppression and no alopecia make X interesting in elderly pts with hormone-refractory disease. Methods: Between Feb 2004 and Oct 2006, 36 elderly (>65 years) MBC pts previously treated with adjuvant therapy and at least one previous hormonal therapy for advanced disease received X 1,000 mg/m2 twice daily, days 1–14 every 3 weeks. Study objectives were to assess efficacy, safety, and impact on quality of life (QoL), assessed by Clinical Benefit Response (CBR) every third cycle. Results: Median age was 70 years (range 68–73), median ECOG PS was 1 (range 0–2). All pts had visceral metastasis and 19 (53%) had ≥2 metastatic sites. A total of 284 cycles were administered (median 6 cycles per pt). After 3 cycles, 10 pts (28%) showed a partial response, 16 (44%) had stable disease (SD), and 2 (6%) had a minor response, resulting in a disease control rate of 78%. Biochemical response (CEA and/or CA 15.3 reduction) was observed in 20 (56%) pts. SD was maintained in 22 pts (61%) after 6 cycles, 10 pts (28%) after 9 cycles and 2 pts (6%) after 12 cycles. Treatment was well tolerated, the most common grade 3 events being mucositis (6%) and hand-foot syndrome (6%). There were no grade 3/4 hematologic toxicities. All adverse events were easily managed with dose adjustments and supportive therapies as required. As a result, all pts (100%) complied with appreciable benefits in terms of QoL (positive CBR in 56% of pts). Conclusions: These preliminary data indicate that this X dose is active and well tolerated in elderly pts with hormone-refractory MBC. This regimen also warrants study as first-line treatment in pts with less aggressive MBC who might not be suitable for combination therapy. No significant financial relationships to disclose.

The alterations induced on the pool sizes of five phospholipid metabolites, glycerol 3-phosphorycholine, glycerol 3-phosphorylethanolamine, phosphorylcholine, sn-glycerol 3-phosphate, and choline were studied by nuclear magnetic resonance (NMR) spectroscopy in murine tumors injected with recombinant murine tumor necrosis factor (TNF). Solid tumors were obtained by s.c. injection of either Friend leukemia cells (clones 3C1-8 and 745) in DBA/2 mice or murine fibrosarcoma cells (HeN4) in C3H/HeN mice. After tumor nodules had developed, TNF or bovine serum albumin was injected intratumorally. Treatment of both tumors with TNF resulted in a marked inhibition of tumor growth. 31P-NMR analyses of Friend leukemia cell tumors (and tissue extracts), 6 h after injection of TNF, showed: (a) a 1.5- to 3.5-fold decrease in the pool sizes of glycerol 3-phosphorylcholine and glycerol 3-phosphorylethanolamine; (b) a 7- to 8-fold increase of sn-glycerol 3-phosphate; (c) a 2- to 3.5-fold decrease of phosphorylcholine; (d) an alkaline shift (0.2 units) in intratumoral pH. Similar metabolic alterations occurred in TNF-treated HeN4 fibrosarcoma. 1H-NMR analyses of Friend leukemia cell tumor extracts also indicated, 6 h after tumor injection with TNF: (a) elevated choline levels (9X); (b) a 19-fold increase in the ratio [choline]/[phosporylcholine]; (c) elevated (1.4X) levels of lactic acid; and (d) a 1.6-fold decrease in the [taurine]/[glycine] ratio. The results are interpreted in the light of possible alterations in the activity of enzymes controlling the de novo biosynthesis and catabolism of phospholipids. We concluded that NMR spectroscopy can be a useful means to monitor the level of some phospholipid precursors and/or derivatives as early markers of therapeutic efficacy in intact neoplastic tissues.

Adult DBA/2 mice were given injections s.c. with either interferon-sensitive (745) or -resistant (3Cl-8) Friend erythroleukemia cells (FLC). After tumor nodules had developed, mouse interferon-alpha/beta was injected daily into the tumor. 31P-Nuclear magnetic resonance (NMR) spectroscopy examinations were undertaken on freshly dissected tumors at different days of treatment with either interferon or control preparations. Analysis of 745 FLC tumors in untreated mice at different days of tumor growth (day 8 to 13 after tumor implantation) showed marked increases in the levels of phosphorylcholine (PCho), glycerophosphorylethanolamine (GroPEtn) and glycerophosphorylcholine (GroPCho). In contrast high levels of PCho, GroPEtn and GroPCho were already detectable in the 3Cl-8 FLC tumors on day 8, and no significant changes were observed during subsequent tumor growth. The intracellular pH value remained practically constant in both FLC tumors. Daily intratumoral administration of either partially purified (10(7) IU/mg of protein) or highly purified (10(9) IU/mg of protein) mouse interferon-alpha/beta to both cell tumors resulted in decreases in the levels of PCho, GroPEtn and GroPCho and in increases in the intracellular pH with respect to tumors treated with control preparations or left untreated. Two days of daily treatment of mice with interferon sufficed to induce these metabolic changes which preceded the appearance of necrosis in the tumors. Treatment of FLC tumors with X-rays on day 12 of tumor growth did not result in any comparable metabolic changes 2 days after irradiation. Changes in the levels of phospholipid metabolites were not observed when 745 or 3Cl-8 cells were cultivated in the presence of interferon. As interferon induced these changes in both interferon-sensitive and -resistant tumors we conclude that interferon treatment results in host-mediated effects on the biosynthesis and/or catabolism of tumor cell phospholipids.

The Enzyme-Linked Immunosorbent assay (ELISA) has been applied to the detection of antibodies to type B influenza virus. The specificity, the sensitivity and the reproducibility of the indirect method has been evaluated. The Haemagglutination inhibition assay (HAI) was used as reference system. The results obtained at a single serum dilution show good correlation between ELISA and HAI test (r = 0.70). Reproducibility is satisfactory. Our results seem to show a better sensitivity of the ELISA test as compared to the HAI test. This feature may be useful for certain applications such as the detection of antibody responses after vaccination. The need of accurate standardization in carrying out the test is pointed out.

The Immunoenzymatic ELISA method has been applied to the detection of antibody to measles virus by using a single serum dilution. The reproducibility has been tested over three groups of sera negative, intermediate and strongly positive, in 48 different assays. The standard deviations and the variation coefficients are reported which show very good reproducibility both within and between runs. As regard sensitivity ELISA has proved more sensitive than Haemagglutination Inhibition (HAI) by testing sera both from healthy and vaccinated individuals. Variation of the absolute O.D. values has been found by using two different lots of plates. The use of control sera with various titres is recommended, and a reference scale may be valuable for normalization of the results.

Murine interferon (IFN) inhibited the growth of 3/3 human tumour xenografts growing in nude mice. This inhibition was independent of the sensitivity of the xenograft to human IFN. In contrast to human IFN, the murine IFN did not directly affect the human tumour cells in vitro as measured by levels of the IFN- induced enzyme 2–5A synthetase or elevation of HLA expression. In preliminary investigations into the role of the host in the murine IFN-mediated tumour inhibition, we found equal activities of this IFN in nude and in beige (NK-deficient) nude mice. Moreover, murine IFN did not elevate levels of NK-cell activity in either of these mouse strains. We conclude that murine IFN mediates this tumour inhibition via an effect on the tumour-bearing host that may or may not involve immune mechanisms.

The sensibility of the Immunoenzymatic ELISA test for the evaluation of antibody response in individuals vaccinated with inactivated Influenza vaccine was evaluated in comparison with the results obtained by the Haemagglutination inhibition test using type B Influenza virus as antigen. The rate of seroconversion obtained with the two methods versus each class of HAI titre or each interval of OD values of the prevaccine sera showed similar features. In 23% of the cases, however, sero-conversion was observed only by the ELISA test, in 5.4% of the cases only by the HAI test. It is to be assessed whether the serological response revealed by the ELISA test is fully correlated with protection against the disease.