1. Inflammosome in the human endometrium: further step in the evaluation of the “maternal side” 
Silvia D'Ippolito, M.D., Chiara Tersigni, M.D., Riccardo Marana, M.D., Ph.D., Fiorella Di Nicuolo, Ph.D., Raffaele Gaglione, M.D., Esther Diana Rossi, M.D., Ph.D., M.I.A.C., Roberta Castellani, L.T., Giovanni Scambia, M.D., Ph.D., Nicoletta Di Simone, M.D., Ph.D. 
Fertility and Sterility 
Volume 105, Issue 1, Pages e4 (January 2016)
DOI: /j.fertnstert
Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

2. Figure 1
NALP-3 and ASC immunohistochemical staining. Representative microphotographs of NALP-3 and ASC staining. Sections (3-μm) of endometrial tissues obtained from 10 fertile women and 27 women with recurrent pregnancy loss were stained with anti-NALP-3 and anti-ASC antibody, as described in Material and Methods section. Endometrial tissues obtained from recurrent pregnancy loss group showed extensive 3+ NALP-3 (B) and ASC (D) staining respect to control group (A and C; original magnification ×200).
Fertility and Sterility  , e4DOI: ( /j.fertnstert )
Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

3. Figure 2
Western blot analysis of NALP-3 and ASC expression in endometrial tissues. Representative Western blot for NALP-3 and ASC protein in endometrial tissue lysates from 10 fertile women, control group (CTR) and 30 women with recurrent pregnancy loss (RPL). β-Actin was used as a loading control. The levels of NALP-3 (A) and ASC (B) protein expression in RPL and CTR groups were evaluated by comparing the constant level of β-actin and expressed as optical density (O.D.). The analysis of total cellular lysates obtained from fertile women and women with RPL showed a significant difference both in NALP-3 (P<.0001) and ASC protein expression (P<.05). ∗Statistical significance vs. control group = P< ξ Statistical significance vs. CTR group = P<.05.
Fertility and Sterility  , e4DOI: ( /j.fertnstert )
Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

4. Figure 3
Caspase-1 activation and interleukin levels. (A) Caspase-1 levels in endometrial tissue lysates obtained from 10 control women (CTR) or 30 women with recurrent pregnancy loss (RPL) were quantified by colorimetric ELISA assay. Results are expressed in as nanograms per milliliter of protein levels. A significant difference between the groups was observed. ∗Statistical significance vs. CTR group = P<.05. (B) Interleukin (IL)-1β levels in endometrial lysates obtained from 10 fertile (control group, CTR) and 30 women with RPL were measured by colorimetric ELISA assay. Results are expressed as nanograms per milliliter of protein levels. ∗Statistical significance vs. CTR = P<.01. (C) IL-18 levels in endometrial lysates obtained from 10 fertile (control group, CTR) and 30 women with RPL were measured by colorimetric ELISA assay. Results are expressed as micrograms per milliliter of protein levels. ∗Statistical significance vs. CTR = P<.01.
Fertility and Sterility  , e4DOI: ( /j.fertnstert )
Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

5. Figure 4
Schematic representation of inflammosome complex pathway. NALP-3 interacts with other intracellular proteins to form a large complex called the inflammasome. At rest, NALP-3 is maintained in an inactive state in the cell cytoplasm. NALP-3 can be activated by a range of pathogen-associated molecular patterns (PAMPs), which include bacterial muramyl dipeptide, bacterial RNA, or viral RNA, and by “danger-associated molecular patterns” (DAMPs) such as ATP, decreased intracellular potassium concentration, increased radical oxygen (ROS) concentration. After activation, NALP-3 interacts with ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain). When associated with the inflammasome, ASC can interact with pro-caspase 1 to mediate its conversion to caspase-1. Caspase-1, in turn, activates the interleukin (IL) precursors pro-IL-1β and pro-IL-18 into their active form IL-1β and IL-18, respectively.
Fertility and Sterility  , e4DOI: ( /j.fertnstert )
Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

6. Supplemental Figure 1
Schematic representation of histopathological analysis of endometrial inflammatory component staining (chronic endometritis, weak or absent inflammatory component) between the recurrent pregnancy loss (RPL) group (27 women) and the control (CTR) group (27 women; P<.0001). CE = chronic endometritis; WFC = weak flogistic stromal component; AFC = absence of flogistic stromal component; P<.0001 vs. CTR.
Fertility and Sterility  , e4DOI: ( /j.fertnstert )
Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

7. Supplemental Figure 2
A semiquantitative immunohystochemical analysis of NALP-3. A Fisher's exact test was performed to determine the statistical difference of NALP-3 immunostaining between the recurrent pregnancy loss group (RPL, 27 women) and the control group (CTR, 10 women). For statistical analysis of immunohystochemistry, the samples were grouped into negative (score, <2) or positive (score, ≥2). We found a significant difference both in stromal and glandular components between CTR and RPL groups (P<.05).
Fertility and Sterility  , e4DOI: ( /j.fertnstert )
Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

8. Supplemental Figure 3
A semiquantitative immunohystochemical analysis of ASC. A Fisher's exact test was performed to determine the statistical difference of ASC immunostaining between the recurrent pregnancy loss group (RPL, 27 women) and the control group (CTR, 10 women). For statistical analysis of immunohystochemistry, the samples were grouped into negative (score, <2) or positive (score, ≥2). We found a significant difference both in stromal and glandular components between CTR and RPL groups (P<.05).
Fertility and Sterility  , e4DOI: ( /j.fertnstert )
Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions