NYU Langone Medical Center New York, NY, United States
Disclosure information not submitted.
Abhimanyu Amarnani1, Doua Azzouz2, MacIntosh Cornwell3, David Mieles3, Peter Izmirly4, Jill Buyon5, Kelly Ruggles6 and Gregg Silverman5, 1NYU Langone Medical Center, New York, NY, 2NYU Langone Health ( School of medicine), New York, NY, 3New York University, New York, NY, 4New York University School of Medicine, New York, NY, 5NYU Grossman School of Medicine, New York, NY, 6NYU Langone, New York, NY
Background/Purpose: Systemic lupus erythematosus (SLE) is associated with gut microbiota dysbiosis, and community imbalance can coincide with flares of Lupus nephritis (LN). Herein, we sought to understand the blood cell gene expression patterns and antibody responses to a gut pathobiont in longitudinal assessments of SLE patients with LN and concurrent gut colonization blooms of Blautia (Ruminococcus) gnavus (RG).
Methods: In a well-characterized SLE cohort, we performed bulk RNA-seq from whole-blood PAXGene tubes. Our cohort consisted of 24 female subjects (8 controls, 8 with non-renal SLE, 4 LN without RG blooms, and 4 LN concurrent with RG blooms) with longitudinal assessment of whole-blood RNA sequencing, lupus activity, and serum anti-RG lipoglycan antibody levels. After quality control and filtering, gene expression profiles were analyzed. Serum levels of the platelet factors, P-selectin and platelet factor 4 (PF4), were assessed by ELISA.
Results: Differential expression and unsupervised clustering analysis revealed distinct gene expression patterns between LN patient without and with RG gut blooms (372 differentially expressed genes; p< 0.01. Gene set enrichment analysis of the differentially expressed genes revealed significant enrichment for “platelet activation”, “platelet alpha granule” and “platelet degranulation” pathways (Figure 1A, FDR < 0.001). Gene expression levels in SLE samples were compared to controls (Figure 1B). As platelet degranulation was found to be upregulated in a subset of LN patients with RG blooms, we interrogated serum levels of P-selectin/CD62P, a protein produced by activated platelets affecting leukocyte activation/migration. We found that P-selectin was increased in LN patients with RG blooms. No significant difference was observed in LN patient without RG blooms or non-renal SLE patients (Figure 2A). The gene expression of PF4, a chemokine protein primary secreted by platelets upon activation was found to be higher in LN RG, compared to controls (Figure 1A). Serum P-selectin directly correlated with PF4 levels (p=0.003)(Figure 2B).
Conclusion: Patients with LN and RG gut blooms displayed upregulated expression of genes for platelet activation and degranulation. Gut blooms with the pathobiont RG, known to cause gut leakiness from the gut and induce high serum anti-RG lipoglycan antibodies, are herein associated with increased platelet activation and raised serum P-selectin and PF4. Taken together, our data point to a potential role for platelet activation and thrombo-inflammation in LN pathogenesis.
Figure 1. LN patients with RG gut blooms overexpress genes, including for platelet activation. (A) Volcano plot of genes differentially expressed in LN patients with (LN RG) and without RG bloom (LN No Bloom) (Red indicates p<0.01 and log2FC>1 or <-1, and green p<0.01 with Log2FC <1 or >-1). (B) Heatmap illustrates differential gene expression in patients with non-renal SLE (SLE Not LN), LN (LN No Bloom), and RG blooms (LN RG), compared to control samples, for 11 genes that intersect with the “Platelet Activation” pathway.
Figure 2. P-selectin levels are increased in patients with LN and RG bloom. (A) P-selectin/CD62-P levels in SLE patients and (B) correlation of P-selectin and PF-4 s in sera of patients with LN and levels of serum RG anti-LG antibodies above level in healthy individuals (p=0.003). *indicates p<0.05, non-parametric Mann-Whitney U analysis of LN RG vs. Control. Based on commercial ELISA of serum (P-selectin 1:400, PF4 1:20, ng/mL).
A. Amarnani: None; D. Azzouz: None; M. Cornwell: None; D. Mieles: None; P. Izmirly: None; J. Buyon: Bristol-Myers Squibb(BMS), 2, GlaxoSmithKlein(GSK), 2, Related Sciences, 1; K. Ruggles: None; G. Silverman: None.
.jpg)
.jpg)
