We would like to acknowledge the excellent technical help of Angela Houwing and help with data collection from Greta Bushnell.

1. Patient Selection
<ol>
	<li>Women with chronic pelvic pain undergo a history and physical exam.</li>
	<li>Localization of pain, timing with regards to the menstrual cycle, associated symptoms including irritable bowel complaints or bladder complaints are noted and recorded.</li>
	<li>Women undergoing laparoscopy are consented for possible resection of endometriosis and use of their tissues for research.</li>
</ol>
2. Laparoscopic Techniques
<ol>
	<li>Laparoscopy is performed using standard techniques through a 5 mm umbilical port and additional 5 mm ports in the lower abdomen.</li>
	<li>Careful inspection of the pelvic is performed to correlate any findings of endometriosis with the pre-operative localization of pain, as indicated by the patient.</li>
	<li>Methylene blue dye is mixed 1:200 with sterile saline and injected onto pelvic surfaces using an aspiration needle.</li>
	<li>Suction irrigation is performed to remove the excess dye and rinse the peritoneal surfaces.</li>
	<li>The pelvis is inspected and the areas of blue dye uptake are recorded.</li>
	<li>When possible, resection of the affected areas of peritoneum is performed.</li>
</ol>
3. Scanning Electron Microscopy
<ol>
	<li>Peritoneal sections are sutured to a sterile telfa board to flatten the specimen and maintain orientation (intraperitoneal side up) and placed in gluteraldehyde fixative (Sigma, St. Louis, MO) for SEM.</li>
	<li>Samples were oriented and post-fixed in 1% buffered osmium tetroxide for 1 hr and dehydrated with sequential ethanol concentrations.</li>
	<li>Samples were dried using liquid carbon dioxide and mounted onto aluminum scanning electron microscope stubs with colloidal silver paste and sputter coated with gold:platinum alloy to a thickness of 20 nM.</li>
	<li>Scanning electron microscopy was performed using a JEOL Model JSM-6400 Scanning microscopy and photomicrographs obtained from 12 random areas.</li>
</ol>

No conflicts of interest declared.

Chronic pelvic pain in women is a costly and poorly understood problem 1. Dysmenorrhea is the most common form of pelvic pain affecting 60% of women 2 and 72% of adolescents 3. Endometriosis is an inflammatory condition that affects 5% of normal women but is present in up to 70% of women with pelvic pain. While endometriosis contributes to chronic pelvic pain (CPP), a mechanism by which lesions cause pain remains uncertain 4. Paradoxically, the severity of pain does not correla...

<table border="1">
 <tbody>
 <tr>
 <td>Name of the reagent</td>
 <td>Company</td>
 <td>Catalogue number</td>
 <td>Comments (optional)</td>
 </tr>
 <tr>
 <td>Methylene blue</td>
 <td>American Reagent</td>
 <td>NDC0517-0310-110</td>
 <td>Mixed 1:200 in saline</td>
 </tr>
 </tbody>
</table>

intraoperative detection of subtle endometriosis: a novel paradigm for detection and treatment of pelvic pain associated with the loss of peritoneal integrity

Endometriosis is a common disease affecting 40 to 70% of reproductive-aged women with chronic pelvic pain (CPP) and/or infertility. The purpose of this study was to demonstrate the use of a blue dye (methylene blue) to stain peritoneal surfaces during laparoscopy (L/S) to detect the loss of peritoneal integrity in patients with pelvic pain and suspected endometriosis. Forty women with CPP and 5 women without pain were evaluated in this pilot study. During L/S, concentrated dye was sprayed onto peritoneal surfaces, then aspirated and rinsed with Lactated Ringers solution. Areas of localized dye uptake were evaluated for the presence of visible endometriotic lesions. Areas of intense peritoneal staining were resected and some fixed in 2.5% buffered gluteraldehyde and examined by scanning (SEM) electron microscopy. Blue dye uptake was more common in women with endometriosis and chronic pelvic pain than controls (85% vs. 40%). Resection of the blue stained areas revealed endometriosis by SEM and loss of peritoneal cell-cell contact compared to normal, non-staining peritoneum. Affected peritoneum was associated with visible endometriotic implants in most but not all patients. Subjective pain relief was reported in 80% of subjects. Based on scanning electron microscopy, we conclude that endometrial cells extend well beyond visible implants of endometriosis and appear to disrupt the underlying mesothelium. Subtle lesions of endometriosis could therefore cause pelvic pain by disruption of peritoneal integrity, allowing menstrual or ovulatory blood and associated pain factors access to underlying sensory nerves. Complete resection of affected peritoneum may provide a better long-term treatment for endometriosis and CPP. This simple technique appears to improve detection of subtle or near invisible endometriosis in women with CPP and minimal visual findings at L/S and may serve to elevate diagnostic accuracy for endometriosis at laparoscopy.

An example of subtle endometriosis that was nearly invisible is shown in Figure 1A. Prior to application of blue dye an irregular area of stippled peritoneum was appreciated aided by reflection of the laparoscopic light. In Figure 1B, the appearance of this same area after staining shows the same irregular pattern over the bladder (Figure 1B). Once this portion of peritoneum was resected (Figure 1C) it was subjected to SEM which showed overlying end...

Watch this Scientific Journal Video about Intraoperative Detection of Subtle Endometriosis: A Novel Paradigm for Detection and Treatment of Pelvic Pain Associated with the Loss of Peritoneal Integrity

Intraoperative Detection of Subtle Endometriosis: A Novel Paradigm for Detection and Treatment of Pelvic Pain Associated with the Loss of Peritoneal Integrity

Loss of peritoneal integrity provides a new paradigm to understand and treat chronic pelvic pain in women with mild forms of endometriosis and can be easily detected using intraoperative instillation of dye at the time of laparoscopy.

Endometriosis is a  common disease affecting 40 to 70% of reproductive-aged women with chronic  pelvic pain (CPP) and/or infertility. The purpose of this ...

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Research

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15.8K Views.  Greenville Hospital System. Loss of peritoneal integrity provides a new paradigm to understand and treat chronic pelvic pain in women with mild forms of endometriosis and can be easily detected using intraoperative instillation of dye at the time of laparoscopy.

Video: Intraoperative Detection of Subtle Endometriosis: A Novel Paradigm for Detection and Treatment of Pelvic Pain Associated with the Loss of Peritoneal Integrity