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Laser capture microdissection and cDNA array analysis of endometrium identify CCL16 and CCL21 as epithelial-derived inflammatory mediators associated with endometriosis.

Chand AL, Murray AS, Jones RL, Hannan NJ, Salamonsen LA, Rombauts L - Reprod. Biol. Endocrinol. (2007)

Bottom Line: Immunoreactive CCL16 was overall higher in glands in ectopic vs. eutopic endometrium from the same woman (P < 0.05).Staining for CCL16 and CCL21 was highly correlated in individual tissues.This study provides novel candidate molecules and suggests a potential local role for CCL16 and CCL21 as mediators contributing to the inflammatory events associated with endometriosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Prince Henry's Institute of Medical Research, Clayton, Victoria, Australia. ashwini.chand@princehenrys.org

ABSTRACT

Background: Understanding the pathophysiology of chemokine secretion in endometriosis may offer a novel area of therapeutic intervention. This study aimed to identify chemokines differentially expressed in epithelial glands in eutopic endometrium from normal women and those with endometriosis, and to establish the expression profiles of key chemokines in endometriotic lesions.

Methods: Laser capture microdissection isolated epithelial glands from endometrial eutopic tissue from women with and without endometriosis in the mid-secretory phase of their menstrual cycles. Gene profiling of the excised glands used a human chemokine and receptor cDNA array. Selected chemokines were further examined using real-time PCR and immunohistochemistry.

Results: 22 chemokine/receptor genes were upregulated and two downregulated in pooled endometrial epithelium of women with endometriosis compared with controls. CCL16 and CCL21 mRNA was confirmed as elevated in some women with endometriosis compared to controls on individual samples. Immunoreactive CCL16 and CCL21 were predominantly confined to glands in eutopic and ectopic endometrium: leukocytes also stained. Immunoreactive CCL16 was overall higher in glands in ectopic vs. eutopic endometrium from the same woman (P < 0.05). Staining for CCL16 and CCL21 was highly correlated in individual tissues.

Conclusion: This study provides novel candidate molecules and suggests a potential local role for CCL16 and CCL21 as mediators contributing to the inflammatory events associated with endometriosis.

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Related in: MedlinePlus

Immunostaining for CCL16 and CCL21 in eutopic and ectopic tissue from control and endometriosis subjects. Staining intensity for (A) CCL16 and (B) CCL21 in glandular epithelium of normal mid secretory endometrium (n = 12) (■) and in glandular epithelium in paired eutopic (▲) and ectopic (▼) endometrium from women with endometriosis (n = 10), also in the mid-secretory phase of the menstrual cycle. Open symbols represent subjects whose samples were used in the gene array study. Staining intensity was scored on a scale of 0 (no stain) to 4 (intense stain). Scores are shown for individual samples. Bars represent mean values. * denotes a statistical difference between 2 samples from the same patients (P = 0.047, Wilcoxon Matched-Pairs Signed-Ranks Test).
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Figure 3: Immunostaining for CCL16 and CCL21 in eutopic and ectopic tissue from control and endometriosis subjects. Staining intensity for (A) CCL16 and (B) CCL21 in glandular epithelium of normal mid secretory endometrium (n = 12) (■) and in glandular epithelium in paired eutopic (▲) and ectopic (▼) endometrium from women with endometriosis (n = 10), also in the mid-secretory phase of the menstrual cycle. Open symbols represent subjects whose samples were used in the gene array study. Staining intensity was scored on a scale of 0 (no stain) to 4 (intense stain). Scores are shown for individual samples. Bars represent mean values. * denotes a statistical difference between 2 samples from the same patients (P = 0.047, Wilcoxon Matched-Pairs Signed-Ranks Test).

Mentions: Since the findings from the cDNA array and real time RT-PCR experiments supported a role for CCL21 and CCL16 in at least some women with endometriosis, immunohistochemistry was performed to establish the presence and cellular location of the protein for these 2 chemokines in eutopic and ectopic endometrial samples. Immunoreactivity for both chemokines was evident in glandular epithelial cells but absent from stromal cells and was overall in accord with the real-time PCR quantitation. CCL16 protein was detected in the glandular epithelium of all but one of the 32 tissues examined and CCL21 protein was evident in glands in all tissues. However, staining was variable between individual tissues even within groups (Figure 3). No significant differences were detected for either CCL16 or CCL21 between mean glandular staining intensity in endometrium from normal women compared with eutopic endometrium from women with endometriosis (Figure 3). The staining pattern in some glands was predominantly apical (Figures 4A–C) suggesting secretion of the chemokine into the uterine lumen. Within each tissue, all glands were equally stained and staining was present in all cells in each gland (Figures 4A–D).


Laser capture microdissection and cDNA array analysis of endometrium identify CCL16 and CCL21 as epithelial-derived inflammatory mediators associated with endometriosis.

Chand AL, Murray AS, Jones RL, Hannan NJ, Salamonsen LA, Rombauts L - Reprod. Biol. Endocrinol. (2007)

Immunostaining for CCL16 and CCL21 in eutopic and ectopic tissue from control and endometriosis subjects. Staining intensity for (A) CCL16 and (B) CCL21 in glandular epithelium of normal mid secretory endometrium (n = 12) (■) and in glandular epithelium in paired eutopic (▲) and ectopic (▼) endometrium from women with endometriosis (n = 10), also in the mid-secretory phase of the menstrual cycle. Open symbols represent subjects whose samples were used in the gene array study. Staining intensity was scored on a scale of 0 (no stain) to 4 (intense stain). Scores are shown for individual samples. Bars represent mean values. * denotes a statistical difference between 2 samples from the same patients (P = 0.047, Wilcoxon Matched-Pairs Signed-Ranks Test).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1884154&req=5

Figure 3: Immunostaining for CCL16 and CCL21 in eutopic and ectopic tissue from control and endometriosis subjects. Staining intensity for (A) CCL16 and (B) CCL21 in glandular epithelium of normal mid secretory endometrium (n = 12) (■) and in glandular epithelium in paired eutopic (▲) and ectopic (▼) endometrium from women with endometriosis (n = 10), also in the mid-secretory phase of the menstrual cycle. Open symbols represent subjects whose samples were used in the gene array study. Staining intensity was scored on a scale of 0 (no stain) to 4 (intense stain). Scores are shown for individual samples. Bars represent mean values. * denotes a statistical difference between 2 samples from the same patients (P = 0.047, Wilcoxon Matched-Pairs Signed-Ranks Test).
Mentions: Since the findings from the cDNA array and real time RT-PCR experiments supported a role for CCL21 and CCL16 in at least some women with endometriosis, immunohistochemistry was performed to establish the presence and cellular location of the protein for these 2 chemokines in eutopic and ectopic endometrial samples. Immunoreactivity for both chemokines was evident in glandular epithelial cells but absent from stromal cells and was overall in accord with the real-time PCR quantitation. CCL16 protein was detected in the glandular epithelium of all but one of the 32 tissues examined and CCL21 protein was evident in glands in all tissues. However, staining was variable between individual tissues even within groups (Figure 3). No significant differences were detected for either CCL16 or CCL21 between mean glandular staining intensity in endometrium from normal women compared with eutopic endometrium from women with endometriosis (Figure 3). The staining pattern in some glands was predominantly apical (Figures 4A–C) suggesting secretion of the chemokine into the uterine lumen. Within each tissue, all glands were equally stained and staining was present in all cells in each gland (Figures 4A–D).

Bottom Line: Immunoreactive CCL16 was overall higher in glands in ectopic vs. eutopic endometrium from the same woman (P < 0.05).Staining for CCL16 and CCL21 was highly correlated in individual tissues.This study provides novel candidate molecules and suggests a potential local role for CCL16 and CCL21 as mediators contributing to the inflammatory events associated with endometriosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Prince Henry's Institute of Medical Research, Clayton, Victoria, Australia. ashwini.chand@princehenrys.org

ABSTRACT

Background: Understanding the pathophysiology of chemokine secretion in endometriosis may offer a novel area of therapeutic intervention. This study aimed to identify chemokines differentially expressed in epithelial glands in eutopic endometrium from normal women and those with endometriosis, and to establish the expression profiles of key chemokines in endometriotic lesions.

Methods: Laser capture microdissection isolated epithelial glands from endometrial eutopic tissue from women with and without endometriosis in the mid-secretory phase of their menstrual cycles. Gene profiling of the excised glands used a human chemokine and receptor cDNA array. Selected chemokines were further examined using real-time PCR and immunohistochemistry.

Results: 22 chemokine/receptor genes were upregulated and two downregulated in pooled endometrial epithelium of women with endometriosis compared with controls. CCL16 and CCL21 mRNA was confirmed as elevated in some women with endometriosis compared to controls on individual samples. Immunoreactive CCL16 and CCL21 were predominantly confined to glands in eutopic and ectopic endometrium: leukocytes also stained. Immunoreactive CCL16 was overall higher in glands in ectopic vs. eutopic endometrium from the same woman (P < 0.05). Staining for CCL16 and CCL21 was highly correlated in individual tissues.

Conclusion: This study provides novel candidate molecules and suggests a potential local role for CCL16 and CCL21 as mediators contributing to the inflammatory events associated with endometriosis.

Show MeSH
Related in: MedlinePlus