Profiling the microbiome of oral and genital mucosal surfaces in Behçet's disease.

Almost 90% of Behçet's patients present with oral and/or genital ulcers which influence the disease outcome. We hypothesised that the dysregulation of the oral and genital microbiome, coupled with dysregulation of the immune response, contributes to the aetiopathogenesis of Behçet's Disease (BD) and drives disease activation. METHOD: 152 BD patient samples; 70 matched oral and genital samples plus 12 unmatched samples (Female: Male, 58:12; mean age, 42 ± 13.9: 39.3 ± 10.3) to profile microbial community high-throughput sequencing of the microbiome using 16 s rRNA sequencing targeting the V1/V2 and V3/V4 hyper variable regions were used and results reviewed in relation to disease severity, Work and Social Adjustment Scale (WSAS) outcomes and medication. RESULTS: Alpha and beta diversity were significantly decreased in genital compared to oral samples; p value<0.05. However, grouping the samples as to whether ulceration was present was not significant. Escherichia-Shigella was the only Amplicon Sequence Variants (ASVs) in the V1/V2 region that was shared between the oral mucosa with ulcer (O_U) and genital mucosa with ulcer (G_U) groups. This was in contrast to the V3/V4 region which indicated that Lachnospiraceae, Saccharimonadales, and Coriobacteriales were shared between the O_U and G_U groups. In addition, gender had no impact on the bacterial abundance in V1/V2 analysis of the oral and genital samples. V3/V4 analysis of genital samples demonstrated that Lactobacilli and Gardnerella were significantly increased in females (20 times) compared to the males in samples; p-adj <0.05. Interestingly in BD patients, Rothia which is commonly found in the mouth was present in both oral and genital samples. Streptococci were significantly increased while Veillonella significantly decreased in the presence of oral ulceration in the BD cohort. The clinical phenotype had no effect on V1/V2 and V3/V4 on the bacterial abundance of oral samples. However, medication e.g. colchicine had a significant effect on the oral microbial abundance (V1/V2; P = 0.020, V3/V4; P = 0.003). There was no relationship between colchicine and the presence/absence of genital ulcers. BD patients with active disease had higher WSAS scores, and their bacterial abundance differed significantly from the non-active BD patients (ADONIS, R2 = 0.05, p value =0.029). CONCLUSION: The presence of the microbes Streptococcus, Veillonella, Gardnerella, Lactobacillus, Atopobium, Peptoniphilus, Corynebacterium and Staphylococcus may provide early evidence of BD patients are with active disease.

Behçet's patients' response to COVID-19 vaccination.

Immune hyperstimulation by SARS-CoV2 results in multi-system involvement with consequent organ damage not dissimilar to Behçet's Disease (BD). Management of BD includes immunosuppressive medication, which led to concerns that; firstly, SARS-CoV-2 would stimulate BD activity, thrombin, clotting times, TPO antibodies, and the effectiveness and duration of the COVID-19 vaccines' response in this potentially vulnerable group. The main objectives of this study were: to assess BD patients' immune response to the COVID-19 vaccines based on age, gender, disease activity, BD phenotype, and immunomodulatory medication compared to healthy control participants by measuring anti-spike IgG levels. Further to evaluate the effect of the COVID-19 vaccines on T and B cells, immunoglobulins, thrombophilia, thyroid function and COVID-19 antibody production. Patients on immunosuppressive medication had a reduced immune response to COVID-19 vaccines. -Also, patients over 40 years and with the neurologic BD phenotype had lower responses. mRNA COVID-19 vaccines were more effective and had fewer side effects compared to conventional COVID-19 vaccines.

Expression of 25-hydroxyvitamin D-1-alpha-hydroxylase, and vitamin D receptor mRNA in normal and malignant breast tissue.

Vitamin D has anti-proliferative and proapoptotic effects on several cell types, including breast cancer cells. There have been no studies quantifying the expression of the enzyme 25-hydroxyvitamin D-1-alpha-hydroxylase (1alphaOHase), which converts 25-hydroxyvitamin D to its active metabolite, in breast tissue. We performed real-time RT-PCR to quantity 1alphaOHase and vitamin D receptor (VDR) mRNA in samples of breast cancer, adjacent non-cancerous tissue and normal breast tissue. 1alphaOHase and VDR mRNA were universally expressed, however, 1alphaOHase expression was significantly down-regulated in adjacent non-cancerous tissue from women with breast cancer in comparison to individuals without cancer. VDR was also up-regulated in breast tumours. The vitamin D axis expression in the breast suggests a role for its actions in normal tissue homeostasis and breast cancer pathogenesis. The decreased expression of 1alphaOHase in normal tissue from women with breast cancer may be important in their predisposition to the development of cancer.

Altered monocyte cyclooxygenase response to lipopolysaccharide in type 1 diabetes.

Type 1 diabetes is caused by adaptive immune responses, but innate immunity is important because monocytes infiltrate islets. Activated monocytes express cyclooxygenase (COX)-2, promoting prostaglandin-E(2) (PGE(2)) secretion, whereas COX-1 expression is constitutive. We aimed to define monocyte COX expression in type 1 diabetes basally and after lipopolysaccharide (LPS) stimulation. Isolated CD14(+) monocytes were analyzed for COX mRNA and protein expression from identical twins (discordant for type 1 diabetes) and control subjects. Basal monocyte COX mRNA, protein expression, and PGE(2) secretion were normal in type 1 diabetic subjects. After LPS, twins and control subjects showed a COX mRNA isoform switch with decreased COX-1 mRNA (P < 0.01), increased COX-2 mRNA (P < 0.01), and increased COX-2 protein expression (P < 0.01). Compared with control subjects, both diabetic and nondiabetic twins showed greater LPS-induced downregulation of monocyte COX-1 mRNA (P = 0.02), reduced upregulation of COX-2 mRNA and protein (P < 0.03), and greater inhibition by the COX-2 inhibitor di-isopropylfluorophosphate (DFP) of monocyte PGE(2) (P < 0.007). We demonstrate an alteration in monocyte COX mRNA expression as well as monocyte COX-2 and PGE(2) production after LPS in type 1 diabetic patients and their nondiabetic twins. Because COX-2 response to LPS is proinflammatory, an inherited reduced response would predispose to chronic inflammatory diseases such as type 1 diabetes.

Vitamin D metabolism in peripheral blood mononuclear cells is influenced by chewing "betel nut" (Areca catechu) and vitamin D status.

CONTEXT: Vitamin D deficiency, common in South Asians, is a risk factor for metabolic syndrome, type 2 diabetes, and ischemic heart disease. Vitamin D receptor (VDR) activation depends on activated vitamin D [1,25-dihydroxyvitamin D (1,25(OH)(2)D)] concentration, reflecting opposing actions of 25-hydroxyvitamin D-1alpha-hydroxylase [1-alpha(OH)ase] for formation and 25(OH)D-24-hydroxylase [24(OH)ase] for catabolism. We previously reported that circulating 1,25(OH)(2)D contributed to determination of VDR-protein levels and VDR genotype was a determinant of both VDR mRNA and VDR-protein in South Asians. OBJECTIVE: We hypothesized that chewing betel nut, an addictive habit common throughout South Asian communities, contributes to hypovitaminosis-D by modulating the enzymes regulating circulating 1,25(OH)(2)D concentration. DESIGN: Peripheral blood mononuclear cell (PBMC) 1-alpha(OH)ase and 24(OH)ase mRNA concentrations were measured and examined in relation to cross-sectional data on the vitamin-D axis, diet, smoking, and betel usage, including PBMC VDR-RNA and VDR-protein content in a pilot study of 33 healthy British Bangladeshis. RESULTS: PBMC 24(OH)ase mRNA correlated positively and serum 1,25(OH)(2)D negatively with betel quids per day (r = 0.49, P = 0.006 and r = -0.486, P = 0.006, respectively). Independent determinants for 24(OH)ase included betel quids per day (P < 0.0001) and serum 25-OHD (P = 0.024). Independent determinants for serum 1,25(OH)(2)D were gender, smoking, and betel quids per day. PBMC 1-alpha(OH)ase mRNA correlated inversely with VDR mRNA (r = -0.44; P = 0.013); its independent determinants were serum 1,25(OH)(2)D and VDR TaqI and BsmI polymorphisms (P = 0.03-0.0001). CONCLUSIONS: Betel chewing is a more powerful independent determinant of increased 24(OH)ase expression and of decreased serum calcitriol than serum 25-OHD, supporting the hypothesis that this habit could aggravate the effects of vitamin-D deficiency.

The mRNA expression of cyclo-oxygenase-2 (COX-2) and vascular endothelial growth factor (VEGF) in human breast cancer.

AIMS: There is a growing body of evidence that cyclo-oxygenase 2 (COX-2) plays an important role in carcinogenesis and angiogenesis of human tumours. The present study aims to compare COX-2 expression in human breast cancer and adjacent non-cancerous tissue (ANCT), and to identify any correlation between COX-2 and VEGF expression. METHODS: Total cellular RNA was extracted from frozen breast tissue samples according to standard methodology. The mRNA copy numbers for COX-2 and vascular endothelial growth factor 189 (VEGF-189) were determined 40 infiltrating carcinomas and 40 matched ANCT specimens using quantitative RT-PCR and TaqMan methodology. RESULTS: The COX-2 mRNA copy number per microg of RNA was two-fold higher in ANCT compared with the cancerous tissue (p = 0.01). Median mRNA copy number was 5.44 x 10(6) for ANCT and 2.30 x 10(6) for tumour, (ANCT range: 1 x 10(6) to 4.12x 10(7)) (tumour range: 1.29 x 10(5) to 1.07 x 10(7)). There was a significant correlation between COX-2 and VEGF-189 mRNA copy numbers in the cancer specimens (correlation coefficient = 0.5528, p = 0.0076). CONCLUSIONS: COX-2 mRNA is overexpressed in both human breast cancer and ANCT. We found higher levels in the matched ANCT which suggests that paracrine effects may be important in the role of COX-2 in mammary carcinogenesis. Furthermore, our results indicate that in human breast cancers COX-2 overexpression is linked to VEGF-189 overexpression and therefore tumour angiogenesis.

Differential expression patterns of the insulin-like growth factor 2 gene in human colorectal cancer.

Tumour development and metastasis are associated with altered gene expression profiles. The aim of this study was to identify the transcriptional differences in normal, tumour and metastatic tissue. We used oligonucleotide arrays to identify differential expression patterns of insulin-like growth factor 2 (IGF 2) between 139 primary colorectal tumour specimens and 42 tumour-adjacent mucosa specimens from colorectal cancer (CRC) patients. The expression levels of the IGF 2 gene were significantly increased in primary tumours compared with adjacent mucosae. This was concordant with our real-time RT-PCR quantification of 48 matched tumour mucosa samples. IGF 2 expression levels were also measured by RT-PCR quantitative analysis in 18 liver metastases and 10 normal tissues from patients without cancer. The mRNA levels were significantly under-expressed in liver metastases compared with either colorectal tumours or adjacent normal mucosae. The non- malignant normal tissue expressed significantly lower IGF 2 levels than adjacent normal tissue, and this was not due to a field effect originating from the tumour. In addition, our microarray data demonstrated that IGF 2 expression was down-regulated in sporadic microsatellite instability (MSI-H) CRC and parallels under-expression of hMLH1 and IGF 2 receptor genes in these patients. We conclude that IGF 2 plays an important role in CRC development. Also, individuals with loss of genomic imprinting (LOI) causing over-expression of IGF 2 may be at greater risk of developing CRC. However, this LOI may be reversed in MSI-H patients.