Plenary Session Abstracts 2021

  • Abstracts are listed in order of presentation
Abstract Number Abstract Title Presenter
3790 Disruption of actin remodeling intensifies aneurysm-promoting signals in smooth muscle cells Madrid
3865 Spleen tyrosine Kinase (Syk): A crucial player of neutrophil infiltration in hepatic IRI Boominathan
3724 Resolution of post-lung transplant ischemia-reperfusion injury is mediated via Resolvin D1-ALX/FPR2 and Maresin 1-LGR6 receptor signaling Leroy
3811 Persistent Immunosuppression and Lack of an Early‚ Cytokine Storm‚ Characterizes COVID-19 as Compared to Bacterial Sepsis Brakenridge
3838 A novel drug combination with improved therapeutic efficacy over standard of care in preclinical models of hepatocellular carcinoma Shrestha
3775 The Stromal Heterogeneity of Pancreatic Cancer and Its Correlation with Patient Survival Tushoski-Alemán
3661 Intestinal microbiota-derived metabolites modulate pancreatic carcinogenesis through tumor infiltrating natural killer cells Yu
3703 Temporal Dynamics of the Intestinal Microbiome Following Short-Term Dietary Protein Restriction Anderson

Surgery Day Abstract Submission 2021 : Entry # 3790
ABSTRACT TITLE
Disruption of actin remodeling intensifies aneurysm-promoting signals in smooth muscle cells
ABSTRACT BODY
Introduction
Mutations within proteins coding for extracellular matrix assembly or those in the transforming growth factor beta (TGFβ) signaling pathway lead to aneurysm formation in patients and mice. As the SMC cytoskeletal network is intricately linked to TGFβ and ECM signaling, we sought to determine if disruption of the cytoskeletal filaments would alter how SMCs respond to angiotensin II, an important mediator of aneurysm formation.
Methods
C57BL/6 mice conditionally lacking Tgbfr1¬ in SMCs (SMC- Tgbr1iko) were generated by crossing Tgfbr1Fl/Fl and Myh11-CreERT2 strains. Jiang et al. have previously shown that these mice develop rapid and severe aortic aneurysm degeneration with 100% penetrance. Analysis was conducted on aneurysm and control mouse and human aortic tissue sections SMCs. α-actin and f-actin were detected by immunofluorescent imaging. The activation status of the NFkB family member RelA and cell growth regulator MAPK (Erk1/2) was assessed by Western blot for phosphoresidues. In other experiments, microtubule and actin stabilizer/destabilizers were employed to determine their effect on NFkB activation in aortic SMCs after AngII stimulation. Cell signaling kinase activities in primary mouse SMCs was assessed via a phosphoantibody array after incubation with AngII and the actin destabilizer cytochalasin B.
Results
We observed that SMC- Tgbr1iko mice had reduced expression of f- and α-actin protein in the aortic media than control mice. Primary Tgbr1iko SMCs also had higher levels of pRelA expression, less α-actin mRNA, and less f-actin than control SMCs. Tissue sections from aneurysm patients had reduced f-actin staining relative to non-aneurysm controls. Disruption of filamentous actin with the destabilizer latrunculin A in vitro led to increased RelA and ERK activation in the presence of AngII, which was further augmented in the presence of a neutralizing antibody for TGFβ. In contrast, jasplakinolide—an actin filament stabilizer—prevented TGFβ blockade induced pRelA expression, suggesting that altered/inhibited TGFβ receptor signaling modulates NFkB through actin depolymerization. While stabilization and disruption of microtubules led to RelA activation, no effect was observed on TGFβ blockade-induced pRelA expression. The phosphoantibody array revealed that f-actin disruption in the presence of AngII activated the CREB, AKT1/2/3, ERK1/2, and STAT3 pathways in mouse aortic SMCs. SMC- Tgbr1iko mice treated with cytochalasin B suffered significantly greater mortality (P=0.031, Log-Rank test) and had increased aortic growth over a 30-day follow-up period.
Conclusions
Aortic aneurysms are coincident with reduced f-actin expression in murine and human aortic SMCs. Chemical deconstruction of actin filaments promotes an inflammatory phenotype in mouse SMCs in vitro, and aggravates aneurysm formation in vivo. Aberrant or inhibited TGFβ signaling potentiates NFkB activation in SMCs, which may be dependent on f-actin disassembly.

PRESENTING AUTHOR NAME
Kyle Madrid
PRESENTING AUTHOR EMAIL ADDRESS
kbmadrid@live.com
PRESENTING AUTHOR ACADEMIC RANK
Other
SENIOR AUTHOR/MENTOR NAME
zhihua Jiang Prof

RESEARCH CATEGORY
Basic Science
RESEARCH DISCIPLINE
Vascular


Surgery Day Abstract Submission 2021 : Entry # 3865
ABSTRACT TITLE
Spleen tyrosine Kinase (Syk): A crucial player of neutrophil infiltration in hepatic IRI
ABSTRACT BODY
Introduction: In orthotopic liver transplantation, hepatic ischemia-reperfusion injury (IRI) plays a significant role, markedly impairing the graft-liver function and increasing graft susceptibility to rejection. Neutrophil activation is a major, but still incompletely characterized event, occurring mainly in the reperfusion phase driving the progression of hepatic IRI. While many of the factors responsible have been identified, no potent pharmacologic treatments have been fully developed. A previously performed multi-omic analysis on post reperfusion liver tissue from a mouse model of hepatic IRI, and human orthotopic liver transplants (OLT), identified SYK as a potential mediator of neutrophil activation IRI. This study was designed to address and understand the underlying mechanism of Syk signaling pathways that are active and their inflammatory process occurring during hepatic IRI. Methods: Adult 10-12 week old mice were split in to two groups: treatment with a) Syk inhibitor Fostamatinib (R788) every day for 5 days prior to IRI, and b) Vehicle everyday for 5 days prior to IRI. Mice in each group were then subdivided in to mice that underwent a 70% partial warm ischemia for 60 minutes, followed by 6 and 24 hours of reperfusion (n=5/group), or mice that were subject only to a sham midline laparotomy (n=3/group). All mice were then euthanized for blood and hepatic tissue collection at each time point. Standard flow cytometry of whole blood was performed with cell specific target markers to identify specific subpopulations. Results: Syk inhibitor Fostamatinib, controlled neutrophil activation and recruitment in hepatic IRI. In mice treated with fostamatinib, the fraction of CD11b+/Ly6G+ neutrophil cells in whole blood after 6h and 24h of IRI was significantly lower, when compared to those treated solely with vehicle. Fostamatinib treatment also reduced the number of activated Ly6G+ neutrophils infiltrating the liver parenchyma at both 6h and 24h post-reperfusion. Furthermore, the inhibition of neutrophil activation was associated with reduced expression of the pro-inflammatory iNOS and increased expression of TGF- at 6h of reperfusion. As a result, mice treated with fostamatinib had significantly reduced serum ALT levels and ameliorated histological hepatic injury at both 6h and 24h of hepatic reperfusion. Conclusion: This study elucidates that preconditioning mice with SYK specific inhibition protects livers from hepatic IRI via the downregulation of neutrophil infiltration. The impact of Syk kinase inhibition in this study corroborates previous potential and new regulatory therapeutic targets for drug therapy in hepatic IRI.

PRESENTING AUTHOR NAME
Vijay Boominathan
PRESENTING AUTHOR EMAIL ADDRESS
vijayboominathan@ufl.edu
PRESENTING AUTHOR ACADEMIC RANK
Postdoctoral (PhD) Trainee
SENIOR AUTHOR/MENTOR NAME
ALi Zarrinpar Dr

RESEARCH CATEGORY
Clinical/Translational
RESEARCH DISCIPLINE
Transplant/Hepatopancreatobiliary (non-cancer)


Surgery Day Abstract Submission 2021 : Entry # 3724
ABSTRACT TITLE
Resolution of post-lung transplant ischemia-reperfusion injury is mediated via Resolvin D1-ALX/FPR2 and Maresin 1-LGR6 receptor signaling
ABSTRACT BODY
Introduction: Dysregulation of inflammation-resolution pathways can lead to post-lung transplant (LTx) ischemia-reperfusion injury (IRI) and allograft dysfunction. We investigated the role of ω-3-derived specialized pro-resolving lipid mediators (SPMs), i.e. Resolvin D1 (RvD1) and Maresin-1 (MaR1), and the respective receptor signaling in the resolution of lung IRI.
Methods: RvD1 and MaR1 expressions were analyzed in bronchoalveolar lavage (BAL) from patients undergoing LTx on post-operative days 0, 1 and 7 using mass spectrometry and ELISA. C57BL/6 mice underwent sham surgery or lung IRI (1hr left lung ischemia followed by 6hr reperfusion) using an in vivo hilar ligation model. RvD1 and/or MaR1 (100ng/kg each; given intratracheally 1hr prior to IR) was administered to mice that were previously treated with/without siRNA for FPR2 (receptor for RvD1) or LGR6 (receptor for MaR1; 10g given i.p. each). Orthotopic left lung transplants were performed between Balb/C brain dead donors and C57Bl/6 recipients to analyze FPR2 and LGR6 receptor expressions at 6hrs and 48hrs post-transplant. Macrophage (MH-S) and epithelial (MLE12) cells were exposed to hypoxia/reoxygenation (HR; 3hr/3hr or 24hrs) followed by cytokine analysis. Groups (n=3-10) were compared using ANOVA with post-hoc Tukey’s test.
Results: RvD1 and MaR1 levels were significantly increased in BAL of LTx patients on day 7 compared to days 0 and 1. In the murine IR model, concomitant treatment with recombinant RvD1 and MaR1 significantly attenuated lung dysfunction after 6hrs compared to either treatment or IR alone, that was abolished by pre-treatment with siRNA for FPR2 and LGR6, respectively. Lung inflammation (IL-17, TNF-, CXCL1, HMGB1), edema (wet/dry ratio) and injury (neutrophil infiltration) were significantly attenuated in mice treated with RvD1+MaR1 compared to either treatment or IR alone. FPR2 and LGR6 expressions were increased in lung tissue after 48hrs compared to 6hrs post-LTx. RvD1 attenuated HMGB1 and TNF- secretion in HR-exposed MH-S cells that was abolished by siRNA for FPR2, and MaR1 treatment mitigated CXCL1 secretion by HR-exposed MLE12 cells that was abolished by siRNA for LGR6.
Conclusions: Our results suggest that resolution of lung IRI is mediated by RvD1-FPR2 and MaR1-LGR6 signaling pathways on macrophages and epithelial cells, respectively. Pharmacological intervention using these specific SPM bioactive isoforms can potentially be used to circumvent pulmonary inflammation and enhance resolution pathways for protection against chronic lung transplant rejection.

PRESENTING AUTHOR NAME
Victoria Leroy
PRESENTING AUTHOR EMAIL ADDRESS
leroyv@ufl.edu
PRESENTING AUTHOR ACADEMIC RANK
Predoctoral (PhD) Trainee
SENIOR AUTHOR/MENTOR NAME
Ashish Sharma PhD, MBBS

RESEARCH CATEGORY
Basic Science
RESEARCH DISCIPLINE
Transplant/Hepatopancreatobiliary (non-cancer)


Surgery Day Abstract Submission 2021 : Entry # 3811
ABSTRACT TITLE
Persistent Immunosuppression and Lack of an Early “Cytokine Storm” Characterizes COVID-19 as Compared to Bacterial Sepsis.
ABSTRACT BODY
Introduction: The presence of hyperinflammation (HI) and early “cytokine storm” has been the presumptive immune dogma surrounding the pathogenesis of COVID-19 infection. However, initial clinical trial results on targeted inflammatory mediator blockade have shown a disappointing lack of efficacy on clinical outcomes. Since superimposed bacterial pneumonia in these patients is common, we hypothesized an immune endotype that included immunosuppression (IS) in COVID-19 infection.

Methods: We enrolled 30 COVID-19, 10 bacterial sepsis, and 10 healthy controls into a UF-CTSI funded pilot prospective observational cohort study. Patient and disease characteristics were collected, as well as blood samples on days 1, 4, 7, 14, 21 and 28 while hospitalized. Plasma immune biomarkers were measured by multiplex immunoassay. We measured peripheral blood CD14+ HLA-DR expression and total myeloid derived suppressor cell (MDSCs, HLA-DRdimCD33+CD11b+) population size and phenotype by flow cytometry. We utilized ELISpot analysis to measure T-cell and monocyte function, as well as responsiveness to ex vivo IL-7 immunostimulant treatment.

Results: The COVID-19 and bacterial sepsis cohorts were similar in patient demographics and comorbidity. However, the sepsis cohort showed significantly worse early physiologic derangement as measured by APACHE II score (12.5 vs 7.5, p=0.03). While maximum SOFA scores were similar, COVID-19 patients had significantly more ventilator days (11.5 vs 3, p=0.02). Compared to controls, both sepsis and COVID-19 patients exhibited elevated inflammatory biomarkers out to 28 days. However, sepsis patients exhibited significantly higher levels of IL-6, IL-1, MCP-1, early after infection (p<0.05) that declined over time. Both COVID-19 and sepsis patients showed evidence of persistent immunosuppression as measured by increased plasma sPDL-1 and decreased HLA-DR expression compared to controls (p<0.01). ELISpot showed a persistent decrease in T-cell IFN-g expression among both COVID and sepsis patients that was improved by ex vivo IL-7 administration. Both COVID-19 and sepsis patients show a significantly elevated percentage of circulating MDSCs among PMBCs compared to healthy controls.

Conclusion: COVID-19 patients exhibit lower early pro-inflammatory biomarkers and physiologic derangement compared to bacterial sepsis patients. Both COVID-19 and sepsis patients show evidence of significantly compromised T-cell immunity after infection, associated with an expansion of circulating MDSCs. Immune endotyping of the individual host response to infection in future precision medicine approach clinical trials is likely a key factor for the successful utilization of immunomodulatory therapy in these diseases.

PRESENTING AUTHOR NAME
Scott Brakenridge
PRESENTING AUTHOR EMAIL ADDRESS
Scott.Brakenridge@surgery.ufl.edu
PRESENTING AUTHOR ACADEMIC RANK
Associate Professor

RESEARCH CATEGORY
Clinical/Translational
RESEARCH DISCIPLINE
Acute Care/Trauma/Sepsis


Surgery Day Abstract Submission 2021 : Entry # 3838
ABSTRACT TITLE
A novel drug combination with improved therapeutic efficacy over standard of care in preclinical models of hepatocellular carcinoma
ABSTRACT BODY
Introduction: Hepatocellular carcinoma (HCC) is the third most common cause of cancer deaths worldwide. Sorafenib, a multi-kinase inhibitor with antiangiogenic and antiproliferative properties, is the preferred chemotherapeutic for HCC, albeit not without a significant incidence of drug resistance and adverse toxicities that can deteriorate patient quality of life. Optimizing drug combinations to exploit synergistic effects may help limit these adverse events while improving survival and efficacy. In-vitro studies in our laboratory employing a phenotype-based precision screening identified 2 of 15 drugs with clinical FDA approval as showing potential for synergistic chemotherapeutic effects in combination with sorafenib in the treatment of HCC. Here, we test the in-vivo efficacy of this 3-drug combination in three independent preclinical models of HCC.
Materials and Methods: 1) C57BL/6 male mice undergoing a 30-week model of NASH driven spontaneous HCC generation were treated orally for the last 8 weeks with vehicle, sorafenib and a 3-drug combination in 10 different dose regimens (n=5-8/gr), sorafenib in regimens varied from 3X to 30X lower than sorafenib alone treatment. 2) Sorafenib, vehicle and 3 regimens of the 3-drug combination were administered to NOD.Cg-PrkdcscidIL2rgtm1Wjl/SzJ (NSG) male mice in 2 distinct subcutaneous HCC cell-line derived xenograft (CDX) models generated by injecting Huh7 and HepG2 cells in their flank (n=5-6/gr). 3) 1 3-drug combination regimen, sorafenib and vehicle were administered to NSG mice in 4 distinct patient-derived HCC subcutaneous xenografts (PDX) models established with flank implantation of tumor biopsies retrieved from 4 patients with HCC undergoing surgical resection at UFHealth (n=3-6/gr) . All mice were euthanized at their respective tumor volume or study endpoint and blood and tissue collected for analysis.
Results: Of the 10 distinct 3-drug combo regimens (R1-R10) used to treat mice undergoing spontaneous NASH-driven HCC, we identified 3 (R2, R4, and R5) that significantly reduced the number of tumor nodules present in mouse livers, as well as both the volume and weight ratios of the tumor nodules, when compared to mice treated with sorafenib alone or vehicle. Mice bearing either Huh7 and HepG2 CDX tumors were sensitive to sorafenib treatment but benefited significantly from the R2 regimen, while R4 and R5 regimens did not demonstrate additive effects on the inhibition of tumor progression. Finally, all PDX tumor models were highly responsive to the R2 regimen, demonstrating significantly more tumor inhibition than higher dose sorafenib monotherapy.
Conclusion: This study provides strong evidence that our phenotype-based precision method efficiently identifies drug combinations to improve therapy. It demonstrates that a novel 3-drug combination with reduced levels of sorafenib has a significant synergistic reduction of tumor burden in independent pre-clinical in vivo HCC models.

PRESENTING AUTHOR NAME
Sunaina Shrestha
PRESENTING AUTHOR EMAIL ADDRESS
sunaina.shrestha@surgery.ufl.edu
PRESENTING AUTHOR ACADEMIC RANK
Biological Scientist
SENIOR AUTHOR/MENTOR NAME
Ali Zarrinpar

RESEARCH CATEGORY
Basic Science
RESEARCH DISCIPLINE
Oncology


Surgery Day Abstract Submission 2021 : Entry # 3775
ABSTRACT TITLE
The Stromal Heterogeneity of Pancreatic Cancer and Its Correlation with Patient Survival
ABSTRACT BODY
Introduction: Pancreatic ductal adenocarcinoma (PDAC) is one of the leading causes of cancer-related death in the United States. Survival outcomes remain dismal despite advances in molecular diagnostics and therapeutics. Recently, intensive research has revealed the remarkable heterogeneity and complexity of the different components in the pancreatic cancer tumor microenvironment (PCTM). However, our knowledge regarding stromal and immune-cell interactions in the PCTM, and how these interactions impact patient outcomes, remains limited. Digital spatial profiling (DSP) technology now allows us to quantitatively analyze transcript and protein expression on intact tissue, offering an opportunity to study the intra-tumor heterogeneity. Here we aim to decipher the stromal factors regulating the tumor-promoting or tumor-restraining components of the PCTM, and their impact on patient outcomes.
Methods: The Nanostring GeoMx DSP is designed for the analysis of high multiplex gene or protein expression on formalin-fixed paraffin-embedded (FFPE) tissue samples. Four FFPE pancreatic tumors from patients with shorter survival (DFS < 12 months, n=2), and longer survival (DFS > 36 months, n=2) were recruited in this study. Prepared slides were stained with immunofluorescent antibodies to facilitate the identification of tissue morphology: Pan-cytokeratin (panCK+) for tumor cells, alpha-smooth muscle actin (aSMA+) for activated myofibroblasts (CAFs), and CD45+ for immune cells. Stained slides were loaded onto a GeoMx instrument and scanned. Six replicated regions of interest (ROI) were annotated from each tissue sample and characterized with 28 protein or 78 RNA probes to determine the expression levels of selected cells within the ROIs. Person’s correlation (R2) was computed for regression of GeoMx DSP data. All statistical analyses were performed using the GraphPad Prism software. P-values were calculated on two-sided t-tests and P < 0.05 was considered statistically significant.
Results: We compared the differential transcriptomes in PDAC cells between the longer and shorter survival groups. Higher expression of antigen presentation gene HLA-E and integrin alpha- and beta- family genes (ITGAV, ITGAX, ITGB2, and ITGB8) was seen in longer-lived patients. These genes are known to regulate immune cell adhesion and migration. Proliferation genes (CCND1 and MKI67) were also found to be increased in activated myofibroblasts in patients with longer survival. Varying abundance and distribution of immune cells within the PCTM were also noted. CD3+ T-cell and CD8+ cytotoxic T-cell infiltration were found to a lesser extent in the tumor site when compared to the non-tumor stromal site.
Conclusions: Our results suggest that further exploration of the protein expression and interactions within the PCTM may improve our understanding of how tumor and stromal heterogeneity impact tumor immunity, and patient outcomes.

PRESENTING AUTHOR NAME
Gerik Tushoski-Alemán
PRESENTING AUTHOR EMAIL ADDRESS
tushoskig@ufl.edu
PRESENTING AUTHOR ACADEMIC RANK
Biological Scientist
SENIOR AUTHOR/MENTOR NAME
Steven Hughes

RESEARCH CATEGORY
Clinical/Translational
RESEARCH DISCIPLINE
Oncology


Surgery Day Abstract Submission 2021 : Entry # 3661
ABSTRACT TITLE
Intestinal microbiota-derived metabolites modulate pancreatic carcinogenesis through tumor infiltrating natural killer cells
ABSTRACT BODY
Introduction: Preclinical data demonstrate that the gut microbiota can promote pancreatic ductal adenocarcinoma (PDAC), but mechanisms remain unclear. We hypothesized that intestinal microbiota alters anti-tumor innate immunity response to facilitate PDAC progression.
Methods: The human PDAC cell line, L3.6pl, was heterotopically implanted into Rag1-/- mice, and the syngeneic murine PDAC cell line, Pan02, was orthotopically implanted into wild type (WT) mouse pancreas. Natural killer (NK) cells were quantitated by flow cytometry. In vivo NK cell depletion was attained by intraperitoneal injection of anti-Asialo-GM1 antibody. Germ-free (GF) and specific pathogen free (SPF) Rag1-/- mice stool was cultured, and bacteria-free (abiotic) supernatant extracted. Cell-free supernatant from a self-assembled 15 bacterial strains were cultured and extracted individually. The ability of these supernatants to regulate human NK-92MI cell cytotoxicity was tested in vitro by flow cytometry. The function of selected bacterial candidates on NK cell activity were then evaluated in vivo. Impact of Enterococcus hirae and NK cells on PDAC tumors was assessed using WT mice.
Results: Compared to GF Rag1-/-, SPF mice developed 292.7% larger PDAC tumors and 64.4% decrease in intratumoral NK cells (p<0.01). These NK cells had 30.9% IFNγ (p<0.01) decrease in SPF-derived tumors vs GF, by flow cytometry. This NK cell associated PDAC phenotype was recapitulated in Rag1-/- mice which had their microbiota depleted with antibiotics vs SPF. Reconstitution of GF Rag1-/- mice with SPF microbiota (ExGF) yielded 138.9% larger tumors (p<0.01) and 78% reduction in NK cell infiltration vs GF mice (p<0.01). Similarly, intratumoral NK cells showed 45.1% decreased IFNγ expression (p<0.01) in ExGF mice compared to GF mice. The anti-tumor effect of microbiota depletion was abrogated by antibody-mediated NK cell depletion resulting in significant increase in PDAC xenograft size in both Rag1-/- and WT mice (p<0.05). Compared to GF, SPF Rag1-/- abiotic stool culture supernatant inhibited NK-92MI cytotoxicity to L3.6pl and its IFNγ expression (p<0.05). Furthermore, NK-92MI exposed to SPF abiotic stool supernatant exhibited reduced expression of mRNA transcripts related to NK cell activation/recruitment and increased expression of inhibitors of anti-tumor pathways as measured by qPCR array. The cell-free supernatant mixture from the 15-strain bacterial consortium replicated the inhibitory impact of Rag1-/- SPF abiotic supernatant with decreased NK-92MI cytotoxicity and IFNγ expression (p<0.05). Commensal bacterial metabolite screen identified E. hirae as an inducer of NK cell cytotoxicity. In vivo studies demonstrated E. hirae inhibits PDAC progression by enhancing intratumoral NK cell activity.
Conclusion: Gut microbiota modulates PDAC tumor progression through NK cell anti-tumor activity. E. hirae enhances intratumoral NK cell activity and inhibits PDAC development in mice.

PRESENTING AUTHOR NAME
Qin Yu
PRESENTING AUTHOR EMAIL ADDRESS
qin.yu@ufl.edu
PRESENTING AUTHOR ACADEMIC RANK
Predoctoral (PhD) Trainee
SENIOR AUTHOR/MENTOR NAME
Ryan M. Thomas

RESEARCH CATEGORY
Basic Science
RESEARCH DISCIPLINE
Oncology


Surgery Day Abstract Submission 2021 : Entry # 3703
ABSTRACT TITLE
Temporal Dynamics of the Intestinal Microbiome Following Short-Term Dietary Protein Restriction
ABSTRACT BODY
Introduction:
A hyperacute inflammatory response in the early days following surgery is associated with enhanced morbidity and mortality and reduced durability of vascular interventions. Opportunities to modulate this response and improve outcomes have been elusive. With established roles in attenuating insulin resistance, oxidative stress, and ischemia-reperfusion injury, short-term (<1 week) caloric and protein restriction (stDPR) is being investigated as a pre-operative conditioning strategy to mitigate surgical stress and improve outcomes. Based on an evolving linkage between the intestinal microbiome and the systemic milieu, we sought to examine the temporal dynamics of the composition and functional phenotype of the intestinal microbiome during stDPR.

Methods:
10 healthy individuals participated in a 4-day diet, individualized to achieve 30% calorie restriction and 70% protein restriction. Participants were permitted to resume an unrestricted diet from days 5-7, and stool samples were collected at baseline, day 4, and day 7. Taxonomic analysis included shotgun metagenomic sequencing (ZymoBIOMICS) followed by prevalence filtering and differential abundance analysis using PIME and ALDEx2 software packages. Functional profiling was performed using HUMAnN 2.0.

Results:
stDPR induced a marked shift in the composition of the microbiome, which almost fully reverted to its baseline configuration within 3 days after resuming an unrestricted diet. 44 species were found to significantly differ (FDR p<0.05) between baseline and Day 4, particularly notable for decreases in butyrate-producing Firmicutes and increases in mucin-degrading Ruminococcus torques. Despite community variations, alpha diversity between conditions failed to show significant changes (Shannon index p=0.24). Accompany the shift in composition, pathway analysis revealed notable changes in metabolic activity between conditions (62 unique MetaCyc pathways, FDR p<0.05). Reductions in amino acid and lipid biosynthesis pathways, as well as increases in catabolic pathways, including nucleotide degradation, were the primary functional changes seen on day 4. Unlike the species composition, however, marked differences in metabolic pathway activity persisted following resumption of an unrestricted diet.

Conclusions:
Notable shifts in taxonomy are seen in as few as four days with stDPR. Interestingly, a relative dysbiosis is present on day 4 with compositional and functional pathway characteristics of inflammatory conditions such as Crohn’s disease, suggesting a stressed state within the intestinal microbiome after stDPR. Similarities in relative abundance before and after dietary intervention reveal a rapid return to baseline. Metabolic characteristics, however, do not fully match this species normalization. Further investigation into the impact of this microbial imbalance is warranted, especially as it relates to systemic stress response pathways.

PRESENTING AUTHOR NAME
Erik Anderson
PRESENTING AUTHOR EMAIL ADDRESS
erik.anderson@surgery.ufl.edu
PRESENTING AUTHOR ACADEMIC RANK
Surgery Resident
Post-Graduate Year (PGY) – for Surgery Residents Only
PGY 4
SENIOR AUTHOR/MENTOR NAME
Scott Berceli

RESEARCH CATEGORY
Clinical/Translational
RESEARCH DISCIPLINE
Vascular


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