Bacterial Translocation Markers as Predictors of Infectious and Inflammatory Complications in Acute Bowel Obstruction

Study Overview
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Brief Summary
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Despite modern approaches to the diagnosis and treatment of acute bowel obstruction (ABO), postoperative mortality ranges from 5 to 32%, and complications occur up 23% of cases. One of the formidable infectious and inflammatory complications of ABO is sepsis. The main component of the development of sepsis in ABO is bacterial translocation (BT). BT is the migration of intestinal bacteria or their products through the intestinal mucosa into the mesenteric lymph nodes and further into normally sterile tissues and organs. Today there are several methods for detecting BT: direct method - the detection of 16s rRNA (ribosomal ribonucleic acid) in mesenteric lymph nodes (MLN); indirect method - the detection of serum lipopolysaccharide-binding protein (LBP) and presepsin (Soluble CD14 subtype or sCD14-ST). The aim of this study is to determine the diagnostic and prognostic significance of bacterial translocation as a predictor of the complications development in patients with malignant and benign acute bowel obstruction by assessing the relationship of biomarkers in the systemic circulation (LBP, sCD14-ST) with the detection of microorganism genes (16s rRNA) in mesenteric lymph nodes.

Detailed Description
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For the early diagnosis of infectious and inflammatory complications, it is necessary to study LBP, sCD-14 and 16sRNA as bacterial translocation markers in patients with malignant and benign acute bowel obstruction, as well as in patients after planned surgical intervention for colon tumors. Based on changes in bacterial translocation biomarkers in the blood serum, it's suggested that patients with researched pathology can be stratified according to the risk level of developing infectious and inflammatory complications. The study materials are blood serum and mesenteric lymph nodes (MLN). Venous blood sampling will be performed 1 hour before surgery, 24 and 72 hours after it. Venous blood will be collected in 5 ml vacutainers with a coagulation activator and a serum gel separator. It will be centrifuged for 20 minutes at 1000 x g, after which the gel completely separates the serum from the clot, forming a tight barrier.ELISA Kit for Lipopolysaccharide Binding Protein (LBP, Human) and for Presepsin (sCD14-ST, Human), from Cloud-Clone Corp. will be used to determine any presence of LBP and sCD14-ST. The analysis will be performed according to the manufacturer's instructions for an ELISA EVOLIS robotic system from BioRad. The operating surgeon will perform a MLN sampling in sterile conditions during surgery after resection of the intestine from the mesentery of the gross specimen. MLN will be placed in a sterile tube without any fillers. The DNA will be extracted by the GeneJET Genomic DNA Purification Kit manufactured by Thermo Fisher Scientific, USA, in accordance with the manufacturer's instructions. The 16s rRNA bacteria in MLN will be detected by using real-time PCR and BIO-RAD CFX96 amplifier with 16s rRNA forward and reverse primers (U16SRT-F FACTCCTACGGGAGGGAGGCAGGT and U16SRT-R TATTACCGCGGCTGCTGGGC). During the implementation, the resources of the Collective Use Laboratory of Research Center Non-profit Joint Stock Company (NJSC) Karaganda Medical University will be used. This research is funded by the Science Committee of the Ministry of Education and Science of the Republic of Kazakhstan (Grant No. AP09260597).

Official Title
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Prognostic Significance of Bacterial Translocation Markers as Predictors of Infectious and Inflammatory Complications in Acute Mechanical Bowel Obstruction

Conditions
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Intestinal Obstruction, Colorectal Cancer, Post-Op Complication, Systemic Inflammatory Response Syndrome, Colon Tumor

Intervention / Treatment
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* Diagnostic Test: LBP
* Diagnostic Test: sCD14-ST
* Diagnostic Test: 16s rRNA


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: patients with malignant acute bowel obstruction, patients with benign acute bowel obstruction, colorectal cancer patients without acute bowel obstruction (planned operations). Exclusion Criteria: age less than 18, pregnancy, patients with paralytic acute bowel obstruction, patients with HIV infection, liver cirrhosis, patient with an infectious process due to another pathology.

Ages Eligible for Study
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Minimum Age: 18 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Malignant ABO<br>60 patients with malignant acute bowel obstruction | Diagnostic Test: LBP<br>* Determine any presence of LBP in blood serum by ELISA method 1 hour before surgery, 24 and 72 hours after it.<br>Diagnostic Test: sCD14-ST<br>* Determine any presence of sCD14-ST in blood serum by ELISA method 1 hour before surgery, 24 and 72 hours after it.<br>Diagnostic Test: 16s rRNA<br>* Determine any presence of 16s rRNA in mesenteric lymph nodes by PCR method.<br>|
| CRC without ABO (control)<br>60 colorectal cancer patients without acute bowel obstruction (planned operations) | Diagnostic Test: LBP<br>* Determine any presence of LBP in blood serum by ELISA method 1 hour before surgery, 24 and 72 hours after it.<br>Diagnostic Test: sCD14-ST<br>* Determine any presence of sCD14-ST in blood serum by ELISA method 1 hour before surgery, 24 and 72 hours after it.<br>Diagnostic Test: 16s rRNA<br>* Determine any presence of 16s rRNA in mesenteric lymph nodes by PCR method.<br>|
| Benign ABO<br>30 patients with benign acute bowel obstruction | Diagnostic Test: LBP<br>* Determine any presence of LBP in blood serum by ELISA method 1 hour before surgery, 24 and 72 hours after it.<br>Diagnostic Test: sCD14-ST<br>* Determine any presence of sCD14-ST in blood serum by ELISA method 1 hour before surgery, 24 and 72 hours after it.<br>Diagnostic Test: 16s rRNA<br>* Determine any presence of 16s rRNA in mesenteric lymph nodes by PCR method.<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Change in number of Participants with Post-operative infectious and inflammatory complications | Аny infectious and inflammatory complications in post-operative period (wound suppuration, anastomotic leak, аbdominal abscesses, peritonitis, sepsis, etc.) | day 3, day 7, day 10 | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| LBP level in serum blood | LBP levels will be compared between groups/ subgroups and in each group/subgroup in dynamic. | 1 hour before surgery, 24 hours after surgery, 72 hours after surgery | 
| sCD14-ST level in serum blood | sCD14-ST levels will be compared between groups/ subgroups and in each group/subgroup in dynamic. | 1 hour before surgery, 24 hours after surgery, 72 hours after surgery | 
| 16s rRNA in mesenteric lymph nodes | Presence or absence of 16s rRNA in mesenteric lymph nodes will be compared between groups/subgroups. | Once (MLN sampling in sterile conditions during surgery) | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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acute bowel obstruction, colon tumor, bacterial translocation, presepsin, lipopolysaccharide-binding protein, 16s rRNA, post-operative complications