"Our previous research has shown OCT to be successful in detecting early lesions in the distal colon of a murine model," researcher Kaitlin Harpel, from the University's BIO5 Institute, told medicalphysicsweb. "In this study, the subsurface view provided by OCT allowed us to detect lymphoid aggregates – clusters of immune cells that play an essential role in the immune response – in the superficial layers of the colon. We are hoping that detection of these structures will provide additional analysis of the tumour microenvironment, leading to a more precise diagnosis of colorectal disease development."

Harpel and colleagues used genetically engineered mouse models to study the relationship between inflammation and cancer. They investigated the effects of inactivating the Smad3 gene (a tumour suppressor) and/or blocking Interleukin 6 (IL-6, a tumour promoter). The study included 24 mice with either knockout (Smad3-/-) or wild-type (Smad3+/+) gene coding for Smad3, and knockout, heterozygous or wild-type gene coding for IL-6 (Il6-/-, Il6+/- and Il6+/+, respectively).

"When the Smad3 gene is inhibited, it can cause chronic intestinal inflammation. This leads to a change in the cell microenvironment, which then triggers accelerated adenoma development," explained Harpel. "The Smad3 knockout model allowed us to monitor the changing microenvironment in the early stages of colorectal cancer tumour development, and analyse how the immune response may be altered in the initial stages of the disease.

OCT imaging

All of the mice were injected with azoxymethane to induce colorectal cancer. The researchers used an endoscopic OCT system (with 8 µm lateral and 4 µm axial resolution) to image morphological changes in the distal colon at 3-week intervals over 12 weeks.

Of 15 mice that completed the study, six developed tumours. At the final time point, OCT showed 100% sensitivity and specificity to nine histologically confirmed lesions. Using OCT, the team calculated a final tumour burden ranging from 0.333 to 31.862 mm3 – results that correlated well with histology. The findings suggest that OCT may prove useful for early detection of colorectal cancer in humans, or for visualizing the effects of treatment.

OCT also successfully detected 61 histologically-confirmed large (above 0.5 mm diameter) lymphoid aggregates in the mucosal and submucosal layers of the colon, with a sensitivity of 87.8% and a specificity of 100%. Sensitivity to aggregates of 0.3 mm or greater was 70.7%, with specificity remaining at 100%. The overall sensitivity of OCT to all 168 lymphoid aggregates (ranging from 0.1 to 2.5 mm diameter) was 49.0%.

Comparing a histological section showing four large and multiple small lymphoid aggregates, with an OCT-generated map confirmed OCT's ability to accurately identify large, but not small, aggregates. "We are now conducting a study with a much higher spatial sampling rate OCT instrument looking at the early stages of adenoma development in azoxymethane-treated mice," noted Harpel. "We are hopeful that this system will make it easier to see the small aggregates to provide a better understanding of how their number or size changes over time."

The researchers found no correlation between the number of aggregates and the number of tumours. However, the median diameter of the lymphoid aggregates in the Smad3−/− group (which had the largest tumour burden) was higher than that of the Smad3+/+ group, suggesting that the size of the aggregates increased with disease severity. They note that more research is required to fully understand the relationship between aggregate size, location and number, and disease development.

At the final time point, the Smad3−/− group had the highest tumour burden, indicating that Smad3 signalling acts as a tumour suppressor. Three groups exhibited zero tumour burden: Il6-/- with either Smad3 type and Il6+/- with Smad3+/+. This finding suggests that agents that inhibit IL-6 could act as novel therapeutics to prevent disease progression and increase the efficacy of anti-cancer agents in colorectal cancer patients.

The researchers also weighed the spleen and thymus at the endpoint, which exhibit inflammation-induced enlargement and shrinkage, respectively. As well as exhibiting the highest tumour burden, the Smad3−/− group had the highest spleen weight and lowest thymus weight, suggesting tumour-induced systemic inflammatory effect on the host immune system.

The authors concluded that, in this limited study, sensitivity and specificity of OCT detection of adenoma and larger lymphoid aggregates was comparable to that of histology, with OCT offering the advantage of being non-destructive. Harpel notes that OCT systems can be easily converted into a hand-held probe, enabling translation of this method from bench to bedside.

"In the future, we hope that OCT will be used in a medical setting to provide earlier detection of colorectal cancer through the subsurface structures it detects as well as the additional microenvironment information that may be gathered through the presence of lymphoid aggregates," she said.

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