A novel method for the analysis of clinical biomarkers to investigate the effect of diet on health in a rat model.

Experiments into the relationship between diet and health have been an area of high interest for a long time. In this study, we investigate the application of multivariate data analysis to differentiate between rat populations fed on two different diets: normal rat diet (control) and Western affluent diet (WAD). Two sets of data were acquired and analysed: one from a biochemical clinical analyser, taking measurements of blood-based biochemical markers; the other from the analysis of the volatile organic compounds (VOCs) emitted from faecal samples from the same animals using selected ion flow tube mass spectrometry (SIFT-MS). Five classes were considered: weanlings, 12 month controls, 12 month WADs, 18 month controls, and 18 month WADs. Data from the biochemical analyser, weanlings and 18 month WAD fed rats showed significant differences from the other measurement classes. This was shown in both the exploratory analysis and through multivariate classification. Classification of control diet versus WAD diets suggested there are differences between classes with 92% accuracy for the 12 month classes and 91% for the 18 month classes. Cholesterol markers, especially as low density lipoprotein-cholesterol (LDL), were the main factor in influencing WAD samples. The data from the SIFT-MS analysis also produced very good classification accuracies. Classification of control diet versus WAD diets using the H3O(+) precursor ion data suggested there are differences between classes with 71% accuracy for the 12 month classes and 100% for the 18 month classes. These findings confirm that total cholesterol and LDL-cholesterol are elevated in the 18 month WAD-fed rats. We therefore suggest that the analysis of VOCs from faecal samples in conjunction with multivariate data analysis may be a useful alternative to blood analysis for the detection of parameters of health.

Multiple sclerosis and COVID-19: Assessing risk perception, patient behaviors and access to disease-modifying therapies.

BACKGROUND: Following the outbreak of COVID-19, global healthcare systems have had to rapidly adapt. People with multiple sclerosis (pwMS) were required to make decisions about their individual risk and consequent work and social behaviors. This study aimed to evaluate risk perception and patterns of shielding behavior amongst pwMS at the onset of the COVID-19 pandemic and the subsequent impact on patients' employment and access to disease modifying therapies (DMTs). METHODS: Postal surveys were sent to 1690 people within a UK population-based MS cohort during the first wave of the COVID-19 pandemic. Patients were surveyed on: (i) perceived vulnerability to COVID-19; (ii) isolation behavior; (iii) interruption to DMT; (iv) employment status; (v) level of satisfaction with their current working arrangement. RESULTS: Responses were received from 1000 pwMS. Two thirds of patients reported isolating at home during the first wave of the pandemic. This behavior was associated with increased age (p<0.0001), higher disability (p<0.0001) and use of high-efficacy DMTs (p = 0.02). The majority of patients reported feeling vulnerable (82%) with perceived vulnerability associated with higher EDSS (p<0.0001) and receiving a high-efficacy DMT (p = 0.04). Clinician-defined risk was associated with shielding behavior, with those at high-risk more likely to self-isolate/shield (p<0.0001). Patients on high-efficacy DMTs were more likely to have an interruption to their treatment (50%) during the first wave of the pandemic. Most pwMS experienced a change to their working environment, and most were satisfied with the adjustments. CONCLUSION: This study highlights the risk perception, social behavioral practices and changes to treatment experienced by pwMS during the first wave of the COVID-19 pandemic in a large, well-described UK cohort. The results may help inform management of pwMS during future pandemic waves.

COVID-19 in Multiple Sclerosis: Clinically reported outcomes from the UK Multiple Sclerosis Register.

BACKGROUND: In March 2020, the United Kingdom Multiple Sclerosis Register (UKMSR) established an electronic case return form, designed collaboratively by MS neurologists, to record data about COVID-19 infections in people with MS (pwMS). OBJECTIVES: Examine how hospital admission and mortality are affected by disability, age and disease modifying treatments (DMTs) in people with Multiple Sclerosis with COVID-19. METHODS: Anonymised data were submitted by clinical teams. Regression models were tested for predictors of hospitalisation and mortality outcomes. Separate analyzes compared the first and second 'waves' of the pandemic. RESULTS: Univariable analysis found hospitalisation and mortality were associated with increasing age, male gender, comorbidities, severe disability, and progressive MS; severe disability showed the highest magnitude of association. Being on a DMT was associated with a small, lower risk. Multivariable analysis found only age and male gender were significant. Post hoc analysis demonstrated that factors were significant for hospitalisation but not mortality. In the second wave, hospitalisation and mortality were lower. Separate models of the first and second wave using age and gender found they had a more important role in the second wave. CONCLUSIONS: Features associated with poor outcome in COVID-19 are similar to other populations and being on a DMT was not found to be associated with adverse outcomes, consistent with smaller studies. Once in hospital, no factors were predictive of mortality. Reassuringly, mortality appears lower in the second wave.

Serum-mediated inhibition of graft-vs.-host reaction.

Rats recovering from a systemic graft-vs.-host reaction (GVHR) possess factors in the serum which can inhibit the production of a local GVHR. After incubation in vitro for 1 h at 37 degrees C these factors reduce the GVH-producing potential of parental spleen or lymph node cells to 24% of control cells treated with normal serum. These factors appear within 1 wk after initiation of a systemic GVHR and some residual activity persists for up to 8 mo. The serum activity was present in the globulin fraction and was completely removed by absorption with spleen, lymph node, or kidney homogenates from either parental strain rats. These studies indicate that during the course of a systemic GVHR, serum factors directed against the host appear in the circulation and tend to inhibit the production of further GVHR by a second challenge of either parental strain cells.

Uptake of 67Ga in the regenerating rat liver and its relationship to lysosomal enzyme activity.

The uptake of 67Ga citrate has been studied in the regenerating rat liver over a period up to 72 hr after partial hepatectomy. The concentration of 67Ga was found to be maximal (four times that of controls) 42 hr after hepatectomy. This was shown to be related to lysosomal enzyme activity rather than to specific phases of the cell cycle, there being a highly significant correlation (p smaller than 0.001) with aryl sulfatase activity. In both regenerating and normal rat livers, it was shown that 67Ga uptake is reduced when protein synthesis is inhibited by cycloheximide but is unaffected by inhibition of DNA synthesis by cytosine arabinoside.

Effects of graft-versus-host reaction on inhibition of tumor growth in vivo and on tumor cytotoxicity in vitro.

In DA X Wistar F1 rats, growth of 10(4) Wistar-specific Sp 1 carcinoma cells s.c. was commonly prevented by a mild subclinical graft=versus-host reaction produced by injecting 50 X 10(6) Wistar spleen cells i.p. either concurrently with the tumor or 7 to 14 days previously, Spleen cells alone had no effect on established tumor, but their injection on Day 14 significantly reduced the recurrence rate after excision of tumor on Day 21. In vitro tests in tumor-bearing rats with graft-versus-host reactions showed increased spleen lymphocyte and serum cytotoxicity; these mechanisms may inhibit tumor growth in vivo. Because Wistar lymphocytes and Sp 1 cells are syngeneic, inhibition of tumor cannot be due to allograft rejection but is probably an effect of increased host immunoreactivity during the graft-versus-host reaction.

Enteral feeding reduces metabolic activity of the intestinal microbiome in Crohn's disease: an observational study.

BACKGROUND/OBJECTIVES: Enteral feeding will induce remission in as many as 80-90% of compliant patients with active Crohn's disease (CD), but its method of action remains uncertain. This study was designed to examine its effects on the colonic microbiome. METHODS/SUBJECTS: Healthy volunteers and patients with CD followed a regimen confined to enteral feeds alone for 1 or 2 weeks, respectively. Chemicals excreted on breath or in faeces were characterised at the start and at the end of the feeding period by gas chromatography/mass spectrometry. RESULTS: One week of feeding in healthy volunteers caused significant changes in stool colour and deterioration in breath odour, together with increased excretion of phenol and indoles on the breath. Feeding for 2 weeks in patients with CD produced significant improvements in symptoms and a decrease in the concentration of C-reactive protein. The faecal concentrations of microbial products, including short-chain fatty acids (SCFAs), and potentially toxic substances, including 1-propanol, 1-butanol and the methyl and ethyl esters of SCFAs, showed significant falls. CONCLUSIONS: A significant change occurs in the production of microbial metabolites after enteral feeding in both healthy volunteers and patients with CD. Many of those detected in CD are toxic and may feasibly lead to the immunological attack on the gut microbiota, which is characteristic of inflammatory bowel disease. The reduction in the production of such metabolites after enteral feeding may be the reason for its effectiveness in CD.

Studies by radioiodination of normal adult, fetal and leukemic cell membranes.

A comparison was made between cord blood lymphocytes, normal adult lymphocytes and leukemic cells after membrane iodination with lactoperoxidase. A double-labeling technique using lactoperoxidase iodination with 125I and 131I followed by analysis on polyacrylamide gel electrophoresis revealed a number of membrane differences between leukemic, normal and fetal cells. There was a reduction in the 70,000 molecular weight component in cord blood cells compared to adult lymphocytes, and an increase in membrane peptides with molecular weights of 35,000, 20,000, 9,000 and 4,000. Although smaller molecular weight peptides were also present in chronic lymphatic leukemia as well as acute myeloid leukemia, these were shown to be distinct from fetal type membrane components.

Factors affecting haemoglobin synthesis in the mouse.

The effect of various forms of haematopoietic stress on haemoglobin production was studied in mice. During acute anaemia, recovery from polycythaemia or irradiation, there was an increase in the number of reticulocytes, and this was associated with a marked increase in the total protein synthesis. The specific activity in reticulocytes increased up to 20-fold from the normal. The maximum specific activity usually preceded the maximum peak of reticulocytosis. The proportion of minor haemoglobin increased by up to 30%, and there was a significant correlation between the specific activity in reticulocytes and the proportion of minor haemoglobin synthesized. These results indicate that erythropoietic stress results in the release of a new population of reticulocytes that are very active in protein synthesis as measured by markedly increased specific activity, and that this coincides with an increase in the proportion of minor haemoglobin produced.

Acridine orange fluorescence cytochemistry for detecting lymphocyte immunoreactivity.

Acridine orange staining reveals changes within 3 hours of in vitro stimulation of normal rat lymphocytes with mitogens, and of immune rat lymphocytes with the sensitizing antigen. An increased number of red fluorescent cytoplasmic organelles, presumably lysosomes are seen by fluorescence microscopy. Fluorimetry of the supernatants from stained cell suspensions suggests an overall decreased cell uptake of the dye. The microscopy and fluorimetry detected early events in the reaction of lymphocytes from tumour-bearing rats with the target tumour cells. It would appear that the changes in intracellular behaviour of the dye and in overall cell uptake after immune stimulation are a reflection of dissociated variations in internal and external cell membrane permeability, and may provide simple general means for recognizing cellular immune reactions.

Progesterone receptor heterogeneity and endometrial tumour differentiation.

Progesterone binding components were isolated using fast protein liquid chromatography (FPLC) with a recovery efficiency of between 60-70%. In normal uterine cytosol, the two components were in about equal proportions. In endometrial cancer, the ratio of peak size between peaks 1 and 2 was 1.5 for Grade I, 3.7 for Grade II and 2.5 for Grade III. Whether estimation of the levels of the two progesterone components separately would enhance predictability of response to hormone therapy in endometrial cancer remains to be established.

Point mutations at codon 12 of C-K-ras in human endometrial carcinomas.

Samples of human endometrial carcinomas of ranging degree of histological differentiation, and normal endometrial tissues were screened for the presence of single site DNA mutations at codon 12 of the C-K-ras gene using oligonucleotide hybridisation of DNA amplified by the polymerase chain reaction (PCR). Two instances of base substitution were detected; one resulting in a GLY-ALA amino acid replacement in a well differentiated (grade 1) tumour and the second causing a GLY-CYS replacement in a poorly differentiated (grade 3 tumour). The small number of samples tested does not permit conclusions to be drawn with respect to correlations between DNA point mutations and the clinical features, age of patient, tumour receptor status and degree of histological differentiation grading.

The role of fetal adrenal hormones in the switch from fetal to adult globin synthesis in the sheep.

The switch from gamma (fetal) to beta (adult) globin production was studied by the analysis of globin synthesis in chronically cannulated ovine fetuses and newborn lambs. The gamma/alpha globin synthesis ratio decreased from 0.98 +/- 0.11 (S.D.) (n = 4 samples) at 100-120 days of gestation to 0.15 +/- 0.07 (n = 4) in lambs of 150-156 days post-conception, and the beta/alpha synthesis ratio increased from 0.04 +/- 0.06 (n = 4) to 1.13 +/- 0.21 (n = 4) over the same period. In bilaterally adrenalectomized fetuses, which survived in utero until 151-156 days, the gamma/alpha and beta/alpha synthesis ratios were 0.64 +/- 0.14 (n = 3) and 0.25 +/- 0.07 (n = 3) respectively in the 150- to 156-day period. Bilateral adrenalectomy did not affect the time of onset of beta globin synthesis, but significantly decreased the rate. In one bilaterally adrenalectomized fetus the infusion of increasing concentrations of cortisol restored the rate of beta globin synthesis to normal. Treatment of three intact fetuses with 100 micrograms cortisol/h for 3 weeks, from 100 to 121 days, did not affect the timing or rate of switch from gamma to beta globin synthesis. Thus fetal adrenal secretions, probably cortisol, affected the rate of change of gamma to beta globin synthesis but other factors must have been involved in the initiation of the switch.

DNA polymorphisms of the human CD46 gene in fetal and adult tissues.

DNA polymorphisms of the CD46 gene were investigated using a full length cDNA probe and five different restriction enzymes in genomic DNA samples from human fetal tissues and adult peripheral blood leukocytes. A HindIII polymorphism was observed and was classed as Types I, II and III at a ratio of approximately 3.5:2:1, depending on the presence or absence of two polymorphic fragments of 4.5 and 2.0 kb. No variation of polymorphic distribution was evident between the fetal and adult samples. Whether variation in CD46 expression between maternal and fetal tissue is relevant to fetal viability in pregnancy remains to be established.

Haemoglobin A2 level in pregnancy.

The haemoglobin A(2) level was measured in 217 pregnant and 137 non-pregnant patients. Twenty-eight patients were reinvestigated at least six weeks after delivery. There was no significant difference in the mean haemoglobin A(2) level in pregnancy (2.4 +/- 0.8%) and the mean haemoglobin A(2) level in the non-pregnant controls (2.3 +/- 0.6%). In all anaemic patients whose A(2) level was raised in pregnancy the level remained elevated after delivery.

Ultrasound-guided fetal blood transfusion for severe rhesus isoimmunization.

Four fetuses with severe rhesus isoimmunization were transfused with packed red blood cells directly into the umbilical vein. The outcome was successful in three. In one infant, this ultrasound-guided technique resulted in resolution of severe fetal hydrops at 27 weeks, allowing delivery of a healthy nonhydropic infant at 33 weeks, and in the other three infants in prolongation of the pregnancy. The last four transfusions were performed after fetal neuromuscular blockade with curare. The procedure would appear to be associated with a low risk of complication and to provide an excellent chance of a successful outcome of a fetus with severe rhesus isoimmunization even when fetal hydrops is present.

The prognostic importance of steroid receptors in endometrial carcinoma.

Three hundred forty-nine cases of primary endometrial carcinoma (endometrioid, adenosquamous, and clear-cell) were studied to investigate the relative prognostic importance of age, menopausal status, stage, histology, myometrial invasion, and estrogen and progesterone receptor content. Excluding menopausal status, all of these variables had a significant relationship to overall survival in a univariate analysis. Using a Cox multivariate regression analysis, stage, age, and an estrogen receptor value of more than 70 fmol/mg protein, combined with a progesterone receptor value of more than 30 fmol/mg protein, were independently associated with survival. The results demonstrate that for maximum prognostic information, both estrogen and progesterone content of tumors should be measured. Maximum prognostic information is obtained by using cutoff levels that are much higher than those traditionally accepted. This has particular relevance for patient stratification in clinical trials investigating receptor information and response to adjuvant or therapeutic treatment.

Increased frequency of autoantibodies in men with sperm antibodies.

Autoantibodies to thyroid microsomes were more frequent in 102 infertile men with complement-dependent sperm-immobilizing activity (sperm immobilization test [SIT] ) in serum (11.8%) than in a control group of 277 men of comparable ages and semen quality without sperm antibodies (4.3%, P less than 0.05). Frequencies of organ-specific antibodies (antigastric parietal cell, antithyroglobulin, and antithyroid microsome) in 57 men with genital tract obstructions and positive SIT were similar to those for control subjects, and there were no significant differences in the frequencies of non-organ-specific autoantibodies (antinuclear antibody, rheumatoid factor, antimitochondrial, and anti-smooth muscle) in the three groups. Because in men without genital tract obstruction antithyroid microsomal autoantibodies were more common with sperm antibodies than without, the possibility of a genetic factor in the causation of sperm autoimmunity should be considered.

The immunoglobulin class of antispermatozoal antibodies in serum.

The aim of this study was to determine the immunoglobulin class of circulating antisperm antibody using a technique called the indirect immunobead test (IBT). In the IBT sperm bound antibodies are detected using polyacrylamide beads coated with rabbit antihuman immunoglobulin classes IgG, IgA, and IgM. Of the 20 infertile men with serum immobilizins, 100% were found to be positive for sperm-bound IgG, 50% positive for IgA, and 0% positive for IgM, using the IBT. Similarly, 20 infertile females with serum immobilizins showed 95% positivity for IgG, 60% for IgA, and 15% for IgM. Thus there was a good correspondence between the presence of serum immobilizins as determined by the sperm immobilization test (SIT) and the IBT. This study provides data that indicates that IgG and IgA are the two major immunoglobulin classes of sperm antibody in male and female immune sera as detected by a simple, sensitive immunological technique, the serum IBT.