Vitamin Supplementation at the Time of Immunization with a Cold-Adapted Influenza Virus Vaccine Corrects Poor Mucosal Antibody Responses in Mice Deficient for Vitamins A and D.

Vitamin A and D deficiencies and insufficiencies are prevalent worldwide in developed and developing countries. Vitamin metabolites are functionally intertwined in that they are high-affinity ligands for related receptors of the nuclear receptor superfamily. The effects of vitamin A deficiencies (VAD) on antibody responses to respiratory virus vaccines have already been demonstrated. Of particular concern was the reduction in IgA, a first line of defense against pathogens in the respiratory tract. Here, we describe the individual and combined effects of vitamin A and D deficiencies in mice immunized with an attenuated influenza virus vaccine. Relative to VAD, vitamin D deficiency (VDD) had a limited effect, but double deficiencies for vitamins A and D (VAD+VDD) further reduced antibody responses in the respiratory tract. The administration of supplemental vitamins A and D to VAD+VDD mice at the time of vaccination restored responses in a dose-dependent manner. Results suggest that vitamin supplementation programs may be beneficial in a clinical setting to promote healthy immune responses to respiratory virus vaccines in vitamin-deficient individuals.

Oral retinyl palmitate or retinoic acid corrects mucosal IgA responses toward an intranasal influenza virus vaccine in vitamin A deficient mice.

Vitamin A deficiency (VAD) is a leading cause of pediatric morbidity and mortality due to infectious diseases. Recent pre-clinical studies have revealed that VAD impairs mucosal IgA-producing antibody forming cell (AFC) responses toward a paramyxovirus vaccine in the upper respiratory tract (URT), thus impeding a first line of defense at the pathogen's point-of-entry. The studies described here tested the hypothesis that VAD may also impair immune responses after FluMist vaccinations. Results show that (i) IgA-producing antibody forming cells (AFCs) are significantly reduced following FluMist vaccination in VAD mice, and (ii) oral doses of either retinyl palmitate or retinoic acid administered on days 0, 3, and 7 relative to vaccination rescue the response. Data encourage the conduct of clinical studies to determine if there are FluMist vaccine weaknesses in human VAD populations and to test corrective supplementation strategies. Improvements in vaccine efficacy may ultimately reduce the morbidity and mortality caused by influenza virus worldwide.

Robust IgA and IgG-producing antibody forming cells in the diffuse-NALT and lungs of Sendai virus-vaccinated cotton rats associate with rapid protection against human parainfluenza virus-type 1.

Sendai virus (SeV), a natural mouse pathogen, shows considerable promise as a candidate vaccine for human parainfluenza virus-type 1 (hPIV-1), and also as a vaccine vector for other serious pathogens of infants including respiratory syncytial virus (RSV). In an effort to define correlates of immunity, we examined the virus-specific serum antibody of cotton rats inoculated intranasally (I.N.) with SeV. Virus-specific antibody forming cells (AFCs) were also measured in the bone marrow, because these are considered responsible for durable serum antibody levels in other viral systems. Results showed that a single SeV inoculation was sufficient to induce virus-specific serum antibodies and bone marrow-resident AFCs that persisted for as many as 8 months post-vaccination. Given that the predominant SeV-specific serum antibody isotype was IgG, an isotype that traffics poorly to the upper respiratory tract (URT), we asked if local nasal and lung-associated antibodies and AFCs were also present. Studies showed that: (i) SeV-specific antibodies appeared in the URT and lower respiratory tract (LRT) within 7 days after immunization, (ii) corresponding AFCs were present in the diffuse-NALT (d-NALT) and lung, (iii) AFCs in the d-NALT and lung peaked at approximately 6 weeks and persisted for the lifetime of the animal, reaching a level exceeding that of the bone marrow by an order of magnitude, (iv) IgA was the dominant isotype among AFCs in the d-NALT and lung at 4-weeks post-vaccination and thereafter, and (v) antibody and AFC responses associated with the prevention of lung infection when animals were challenged with hPIV-1 just 1 week after vaccination.

Respiratory vaccination of mice against influenza virus: dissection of T- and B-cell priming functions.

We find that a single respiratory administration of replicationally inactivated influenza A viral particles most often elicits a waning serum antibody response, as the long-sustained bone marrow antiviral plasma cell populations characteristically induced by viral infection are lacking, though antiviral plasma cells at other sites may occasionally persist for a long time. To determine whether this alteration in the pattern of the B-cell response is a reflection of the nature of T-helper (Th) priming, we simultaneously primed B cells with inactivated influenza A/PR8(H1N1) and Th cells with infectious A/x31(H3N2). We show that Th cells cross-react extensively between these two viruses, although the antibody response to viral envelope glycoproteins is completely non-cross-reactive. Th cells primed by infectious A/x31 have little impact on the antibody response specifically elicted from naïve B cells by inactivated A/PR8 viruses, suggesting that the characteristic vigour of the antibody response to influenza viral infection depends on the direct interaction of antiviral B cells with virally infected dendritic cells. Memory B cells primed by inactivated influenza viral particles however, respond rapidly to secondary challenge with live or inactivated viruses, promptly populating bone marrow with antiviral plasma cells. Moreover, Th cells primed by previous live A/x31 viral challenge alter the pattern of the response of naïve B cells to live A/PR8 challenge by accelerating the appearance of anti-H1/N1 plasma cells in bone marrow, eliminating the early spike of anti-H1/N1 plasma cells in the mediastinal node, and generally diminishing the magnitude of the lymph node response. Inactivated A/PR8 and infectious A/x31 are both effective vaccines against A/PR8 infection, as mice preimmunized with either vaccine exhibit much more rapid viral clearance from the lung after infectious A/PR8 challenge. In fact, even when given during a course of anti-CD8 treatment to preempt cross-reactive cytotoxic T cells, live A/x31 is a more effective vaccine against A/PR8 infection than is inactivated A/PR8 itself.

Long lived multi-isotype anti-HIV antibody responses following a prime-double boost immunization strategy.

Despite decades of work, an effective HIV vaccine remains elusive. In an effort to elicit protective immunity, investigators have sought to define vaccines able to elicit durable HIV-specific B-cell and T-cell activities. Additionally, vaccines are sought which can induce antibodies of a variety of isotypes, as each isotype possesses unique attributes in terms of opsonization, Fc receptor binding capacity, complement fixation and location. One prominent new vaccine strategy, applied to numerous distinct antigenic systems is the prime boost-regimen, with DNA, vaccinia virus (VV), and/or purified recombinant protein. To examine the durability, location and isotype distribution of responses induced by prime-boost regimens, we tested successive immunizations with DNA, VV and protein (D-V-P), comparing three forms of protein inoculations: (i) purified protein administered intramuscularly with complete Freunds adjuvant, (ii) purified protein administered intranasally, and (iii) purified protein conjugated to oxidized mannan, administered intranasally. We found that all three protocols elicited serum antibodies of multiple isotypes, with serum IgA being most prominent among mice immunized with mannan-conjugated protein. All D-V-P protocols, regardless of protein form or route, also elicited antibody responses at mucosal surfaces. In bronchoalveolar lavage, a tendency toward IgA production was again most prominent in mice boosted with the protein-mannan conjugate. Both B-cell and T-cell responses were sustained for more than 1 year post-immunization following each form of vaccination. Contemporaneous with long-lasting serum and mucosal antibodies were antibody forming cells in the bone marrow of primed animals. Results highlight the D-V-P vaccination strategy as a promising approach for attaining durable, multi-isotype B-cell and T-cell activities toward HIV.

Localized whole eye radiotherapy for retinoblastoma using a (125)I applicator, "claws".

PURPOSE: To treat children with retinoblastoma, who require whole eye radiotherapy, with a specially designed (125)I applicator that irradiates the eye while sparing the surrounding tissues. METHODS AND MATERIALS: Under general anesthesia, a pericorneal ring is attached to the 4 extraocular muscles, and 4 appendages, each loaded with (125)I seeds, are inserted beneath the conjunctiva in-between each pair of muscles and attached anteriorly to the ring. Twenty-nine eyes were treated. Eighteen received a median dose of 28 Gy during 91 hours and 11 received 40 Gy during 122 hours, when the relative biologic effectiveness was taken as 1 instead of 1.5. Six had received prior chemotherapy. RESULTS: Twenty-four eyes were followed up for 2-157 months (median 29). Although 22 eyes responded, local control was achieved in 13 patients, 3 of whom required additional treatment for new tumors; a further 3 required additional treatment for tumor recurrence as well as new tumors. One of these eyes was enucleated for neovascular glaucoma. All 6 Group I-III eyes and 6 of 18 Group V eyes were retained for 2-157 months (median 39), with good vision in 10 eyes. Three developed cataracts 7, 8, and 12 years later, 1 of which has been removed. CONCLUSIONS: This is a new way of irradiating the whole eye with a minimal dose to the surrounding tissues. The treatment time is only 5 days. It is effective in Groups I-III, but only 33% of Group V eyes retained vision. No late cosmetic defects occurred.

TH cells primed during influenza virus infection provide help for qualitatively distinct antibody responses to subsequent immunization.

The quality of the primary Ab-forming cell (AFC) response in cervical lymph nodes and mediastinal lymph nodes of mice to intranasal influenza virus was strongly influenced by viral replicative capacity. IgA secretors were prominent in the early AFC response to infectious virus in mediastinal lymph nodes, while IgG expression was more frequent among isotypically switched AFC in cervical lymph nodes of the same mice; this pattern was reversed in the response to inactivated virus. Influenza viruses A/Puerto Rico/8/34 (A/PR8) and A/X-31 share six of eight genome segments, differing only in hemagglutinin (H1 in A/PR8, H3 in A/X-31) and neuraminidase (N1 in A/PR8, N2 in A/X-31) genes. These viruses therefore elicit extensively cross-reactive TH populations, though their glycoproteins are serologically unrelated. Mice recovered from an A/X-31 infection thus mount a primary B cell response against A/PR8 glycoproteins, when challenged with the latter virus, though this response can call upon memory TH cells. To assess the impact of memory TH populations on a primary Ab response, we compared the AFC response to inactivated A/PR8 in naive mice and mice that had cleared an A/X-31 infection. A/X-31 immune mice mounted a more vigorous AFC response against A/PR8 H1 and N1 glycoproteins than naive animals, when immunized intranasally with inactivated A/PR8. However the distribution of isotypes among H1/N1-specific AFC in lymph nodes of A/X-31-primed mice resembled that of naive mice. Evidently, in this functional context, memory TH cells retained the ability to help Ab responses different in quality from that generated during their primary reaction.

Carcinoma of the cervix and the use of hyperbaric oxygen with radiotherapy: a report of a randomised controlled trial.

BACKGROUND AND PURPOSE: A randomised controlled trial of hyperbaric oxygen in the radiotherapy of Stage IIb and III carcinoma of cervix was performed between 1971 and 1980. Apart from an abstract giving an interim report in 1977, results have not been published. MATERIAL AND METHODS: In a four arm study, 335 patients were randomised to treatment in 10 or 28 fractions, in hyperbaric oxygen or in air. Data is available concerning 327 cases and this has been analysed. RESULTS: There was no advantage in tumour control shown with the use of hyperbaric oxygen. There was evidence for an increase in late radiation morbidity when treatment was given in hyperbaric oxygen rather than in air and when, using 10 fractions, a total dose of 45 rather than 40 Gy was achieved. For late intestinal morbidity, the fractionation sensitivity (alpha/beta ratio) was calculated to be 4.3 Gy and the steepness of the dose response curve (gamma50) to be 2.6. CONCLUSIONS: Hyperbaric oxygen gave no benefit in the treatment of patients with stage IIb and III carcinoma of the cervix treated with radiotherapy using two fractionation regimes. Important data regarding late radiation morbidity has been revealed.

Matching antibody class with pathogen type and portal of entry: cognate mechanisms regulate local isotype expression patterns in lymph nodes draining the respiratory tract of mice inoculated with respiratory viruses, according to virus replication competence and site of inoculation.

Intranasal deposition of Sendai virus (SV) in C57BL/6 mice provokes an Ab-forming cell (AFC) reaction in mediastinal (MLN) and cervical lymph nodes (CLN), which drain the lungs and upper respiratory tract, respectively. While the majority of AFC elicited by infectious SV at both sites produced IgG, the CLN response to SV rendered inactive in replication was restricted almost entirely to IgA, although isotype switching in mediastinal continued to be skewed heavily to IgG. However, in vitro restimulation of the accompanying virus-specific T cell populations from the two sites did not reveal any significant difference in lymphokine output, and isotype expression was not altered substantially in mice lacking IL-4 or IL-6 genes. To dissociate the response to specific Ags from the inflammatory reaction to viral infection, we examined the response to inactivated SV in the face of infection with influenza virus A/HKx31. The magnitude and IgA dominance of the anti-SV AFC population in the CLN were unaffected by a simultaneous, vigorous, IgG-dominated CLN anti-influenza reaction. Evidently, the characteristics of this antiviral response are determined primarily by cognate interactions. Moreover, the IgA bias of the CLN AFC response to inactivated SV was observed only when the virus was delivered intranasally: injection under the epidermis of the cheek, a site that has a lymphatic drainage into the CLN, resulted in an IgG-dominated CLN AFC reaction, lacking IgA. The site of deposition of a vaccine can thus have more influence on the pattern of isotypes induced than the site at which the immune response is initiated.

Intranasal Sendai virus vaccine protects African green monkeys from infection with human parainfluenza virus-type one.

Human parainfluenza virus-type I (hPIV-1) infections are a common cause of "group" and hospitalizations among young children. Here we address the possibility of using the xenotropic Sendai virus [a mouse parainfluenza virus (PIV)] as a vaccine for hPIV-1. Sendai virus was administered to six African green monkeys (Cercopithecus aethiops) by the intranasal (i.n.) route. A long lasting virus-specific antibody response was elicited, both in the serum and nasal cavity. Sendai virus caused no apparent clinical symptoms in the primates, but live virus was detected in the nasal cavity for several days after inoculation. No virus was detected after a second dose of Sendai virus was administered on day 126 after the initial priming. Animals were challenged with hPIV-1 i.n. on day 154. All six vaccinated animals were fully protected from infection while six of six control animals were infected with hPIV-1. The antibody responses induced by Sendai virus immunizations proved to be greater than those induced by hPIV-1. These results demonstrate that unmanipulated Sendai virus is an effective vaccine against hPIV-1 in a primate model and may constitute a practical vaccine for human use.

Distinctive kinetics of the antibody-forming cell response to Sendai virus infection of mice in different anatomical compartments.

The single-cell ELISPOT assay was used to determine the frequency and isotype commitment of virus-specific antibody-forming cells (AFC) at different anatomical locations following intranasal Sendai virus infection of C57BL/6 and 129/Sv mice. AFC responses in the mediastinal and cervical lymph nodes showed sharp increases and declines, first of IgM AFC, peaking about 7 days after infection, and then of IgG and IgA AFC, peaking about 10 days after infection. A wave of IgM AFC preceding the other isotypes was less evident in the spleen, where peak frequencies of AFC occurred 14 days after infection. Virus-specific AFC appeared in the bone marrow with a unique kinetic pattern, increasing in frequency gradually over the first 3 weeks after infection to a plateau that remained constant. Circulating IgM and IgG achieved significant titers approximately a week after infection; IgM titers were transient, but IgG levels increased sharply and remained high, reflecting the longevity of the bone marrow AFC response. Strain differences in isotype bias were noted, particularly preferential switching to the gamma 2a gene in 129/Sv mice. The B-cell response to acute respiratory viral infection thus exhibits features that are distinct from the primary response to nonreplicating antigens.

Respiratory virus infection of mice provokes a permanent humoral immune response.

We have observed that respiratory virus infection of mice provokes an extremely persistent humoral immune reaction, due to a long-sustained population of antibody-secreting cells in the bone marrow. Theories of humoral immunity that strongly distinguish primary and secondary reactions thus may not adequately describe the immune response to respiratory viruses.

Improved iodine-125 plaque design in the treatment of choroidal malignant melanoma.

The use and development of iodine-125 plaque therapy for choroidal malignant melanoma are described. Since 1975 experience has led to changes in plaque design and insertion techniques. Twenty-one patients were irradiated with local episcleral iodine-125 plaques. Three patients required a second plaque for tumour recurrence. Four eyes were enucleated because of continued tumour growth and a further eye was removed because of glaucoma secondary to radiation retinopathy. Two patients (9.5%) died of metastases. The remaining 19 patients are alive and clinically clear of metastases, with a mean follow up time of 73.1 months (range 43-142 months).

A dominant idiotype in the antibody response against the influenza virus hemagglutinin. Serum and in situ analyses.

PY206 is an Id associated with a BALB/c murine mAb described as being specific for the influenza A virus hemagglutinin. However, production of this Id by BALB/c mice immunized with influenza is low. This report shows that the PY206 Id is a dominant component of the anti-influenza antibody response in C57BL/6J strain mice infected intranasally with the influenza A/Hong Kong/168/(H3N2)[R] X-31 virus. High PY206 Id expression was linked to the IgHb Ig allotype locus. PY206 Id+ antibody-forming cells were identified in situ in cryostat sections of lymphoid tissues and idiotypic heterogeneity was identified among PY206+ B cells. Uninfected adult C57BL/6J mice had PY206 Id in their serum that lacked influenza binding specificity. In situ analysis of prenatal and neonatal spleen of uninfected C57BL/6J mice showed that the expansion of PY206 Id+ B cells occurred early in development. PY206+ cells were demonstrated in the lungs of influenza-infected mice but not in normal mice, establishing the capability to study this B cell population in the lung. This model offers the opportunity to manipulate the anti-influenza A virus hemagglutinin B cell response and to study the proliferation and migration of influenza-specific B cells in their native tissue environments.

Hyperbaric oxygen in the radiation treatment of head and neck cancers.

Published reports on the use of hyperbaric oxygen in the radiation therapy of head and neck cancer are reviewed. The weight of evidence, from randomised trials, is that it is of clinical value in the control of medium sized head and neck tumours. It is also of value in the treatment of lymph node disease. Four attempts to improve the results of the treatment, by various physiological measures, are reviewed. It is concluded that hyperbaric oxygen may be of use when combined with nitroimidazoles as a radiation sensitiser and warrants further study when combined with induced anaemia, aimed to reduce the tumour cord and prevent repopulation between fractions.

The treatment of tumors by the induction of anemia and irradiation in hyperbaric oxygen.

Because increased effects have been achieved when murine tumors are irradiated after a period of hypoxia and because of anecdotal clinical experiences of an improved result after irradiation of previously anemic patients in hyperbaric oxygen, the relationship between irradiation and increased survival was investigated in seventy-two patients with advanced head and neck or cervical cancer. Anemia was achieved by means of a two-stage isovolemic venesection maintained for seventy-two hours, hemoglobin was returned to a normal level, and treatment in hyperbaric oxygen was started. Marked tumor shrinkage after the induction of anemia and before radiotherapy was seen and was probably disease, site, and hemoglobin level related. As a result, a possible new approach to cancer therapy is suggested. After completion of therapy, the 1-year disease-free survival for patients with head and neck and cervical cancer was not improved, but the 21-month survival for cervical cancer was improved. Further studies are strongly urged.