Characterization of tubal occlusion after transcervical polidocanol foam (PF) infusion in baboons.

OBJECTIVE: Our long-term goal is to develop a nonsurgical method of fallopian tubal occlusion for the purpose of permanent contraception. We have previously demonstrated that transcervical administration of 5% polidocanol foam (PF) can create tubal occlusion in macaques but that multiple treatments are required. In this study, we assessed the efficacy of various regimens of PF with and without depomedroxyprogesterone acetate (DMPA) (to control ovarian cycle phase) in the baboon. STUDY DESIGN: Adult cycling female baboons were evaluated for tubal patency by hysterosalpingography and then received a transcervical infusion of PF with (+) or without (-) an intramuscular injection of DMPA (3.5 mg/kg). Two concentrations of PF were compared: 1% [(+) DMPA, n=5; (-) DMPA, n=3] and 5% [(+) DMPA, n=4; (-) DMPA, n=3]. Controls received (+) DMPA (n=2) or (-) DMPA, (n=3) only. The reproductive tracts were removed 1-3 months after treatment for examination. RESULTS: No fallopian tubal occlusion was observed in negative controls (±DMPA). Histologic complete tubal occlusion was observed in 3/8 of females treated with 1% PF and in 6/7 treated with 5% PF. Histologic evaluation suggested that 1% PF is associated with prolonged chronic inflammation (more than 2-3 months), while 5% treatment eliminates the epithelial lining, at least focally, and resolves into complete occlusion within 1-2 months. This pattern of complete occlusion was seen in all 4 females that received 5% PF (+DMPA) and in 2/3 that received 5% PF (-DMPA). CONCLUSION: In a baboon model of transcervical permanent contraception, a single treatment with 5% PF resulted in complete tubal occlusion more reliably (85%) than 1% PF (38%). Cotreatment with DMPA may improve treatment results with 5% PF but requires additional study. IMPLICATIONS: A finding that a single transcervical treatment with 5% PF can occlude the fallopian tubes of baboon supports further study of this approach as a novel strategy for permanent contraception for women.

Pathogenesis of fallopian tube damage caused by Chlamydia trachomatis infections.

Chlamydia trachomatis is the leading cause of bacterial sexually transmitted disease worldwide resulting in 4-5 million new cases of Chlamydia annually and an estimated 100 million cases per annum. Infections of the lower female genital tract (FGT) frequently are asymptomatic; thus, they often remain undiagnosed or untreated. If infections are either not resolved or left untreated, chlamydia can ascend to the upper FGT and infect the fallopian tubes (FTs) causing salpingitis that may lead to functional damage of the FTs and tubal factor infertility (TFI). Clinical observations and experimental data have indicated a role for antibodies against C. trachomatis proteins such as the 60-kDa heat shock protein 60 (cHSP60) in the immunopathogenesis of TFI. When released from infected cells, cHSP60 can induce proinflammatory immune responses that may functionally impair the FTs leading to fibrosis and luminal occlusion. Chlamydial pathogenesis of irreversible and permanent tubal damage is a consequence of innate and adaptive host immune responses to ongoing or repeated infections. The extracellular matrix that is regulated by metalloproteinases may also be modified by chlamydial infections of the FGT. This review will highlight protective and pathogenic immune responses to ongoing and repeated chlamydial infections of the FGT. It will also present two recent hypotheses to explain mechanisms that may contribute to FT damage during a C. trachomatis infection. If Chlamydia immunopathology can be controlled, it might yield a method of inducing fibrosis and thus provide a means of nonsurgical permanent contraception for women.

Presence of hydrosalpinx correlated to endometrial inflammatory response in vivo.

OBJECTIVE: To evaluate and compare the inflammatory response and mediators in the endometrium of patients with hydrosalpinges compared with normal controls. DESIGN: Retrospective case-control study. SETTING: Urban medical center. PATIENT(S) AND INTERVENTION(S): Hysterectomy samples were identified as being affected by hydrosalpinx or salpingitis (n = 30) and were age-matched with control samples (n = 30). INTERVENTIONS: Fallopian tube and endometrial slides were analyzed for leukocytes and immunohistochemical techniques performed for cytokines (interleukin-2 [IL-2]). MAIN OUTCOME MEASURE(S): Evaluate and compare the endometrial inflammatory response (leukocytes and cytokines) from samples affected and non-affected by hydrosalpinx and salpingitis. RESULT(S): Examination of tubal and endometrial slides with hydrosalpinx demonstrated a statistically significant increase in the number of overall inflammatory cells. High-intensity immunohistochemical staining for IL-2 was demonstrated in 7.4% of controls versus 65% of cases. CONCLUSION(S): A defined, identifiable, local response to hydrosalpingeal fluid has been demonstrated in the endometrium. This response consists of statistically significant elevations of leukocytes and IL-2. An inflammatory endometrial response may be an independent contributor to the decreased reproductive outcome observed in patients with hydrosalpinges.

Infection of human fallopian tube epithelial cells with Neisseria gonorrhoeae protects cells from tumor necrosis factor alpha-induced apoptosis.

Following infection with Neisseria gonorrhoeae, bacteria may ascend into the Fallopian tubes (FT) and induce salpingitis, a major cause of infertility. In the FT, interactions between mucosal epithelial cells and gonococci are pivotal events in the pathogen's infection cycle and the inflammatory response. In the current study, primary FT epithelial cells were infected in vitro with different multiplicities of infection (MOI) of Pil+ Opa+ gonococci. Bacteria showed a dose-dependent association with cells and induced the secretion of tumor necrosis factor alpha (TNF-alpha). A significant finding was that gonococcal infection (MOI = 1) induced apoptosis in approximately 30% of cells, whereas increasing numbers of bacteria (MOI = 10 to 100) did not induce apoptosis. Apoptosis was observed in only 11% of cells with associated bacteria, whereas >84% of cells with no adherent bacteria were apoptotic. TNF-alpha was a key contributor to apoptosis, since (i) culture supernatants from cells infected with gonococci (MOI = 1) induced apoptosis in naïve cultures, suggesting that a soluble factor was responsible; (ii) gonococcal infection-induced apoptosis was inhibited with anti-TNF-alpha antibodies; and (iii) the addition of exogenous TNF-alpha induced apoptosis, which was inhibited by the presence of increasing numbers of bacteria (MOI = 10 to 100). These data suggest that TNF-alpha-mediated apoptosis of FT epithelial cells is likely a primary host defense mechanism to prevent pathogen colonization. However, epithelial cell-associated gonococci have evolved a mechanism to protect the cells from undergoing TNF-alpha-mediated apoptosis, and this modulation of the host innate response may contribute to establishment of infection. Understanding the antiapoptotic mechanisms used by Neisseria gonorrhoeae will inform the pathogenesis of salpingitis and could suggest new intervention strategies for prevention and treatment of the disease.

TNF contributes to the immunopathology of perforin/Fas ligand double deficiency.

Perforin (pfp)/Fas ligand (FasL) double-deficient mice have previously been shown to be infertile, lose weight and die prematurely due to tissue destruction caused by a significant inflammatory infiltrate of monocytes/macrophages and T cells. Herein we have compared disease progression in mice additionally deficient in the inflammatory mediator TNF. Unlike pfp/FasL double-deficient mice (TNF+/+ pfp-/- gld), mice lacking functional TNF, FasL and pfp (TNF-/- pfp-/- gld) were comparatively fertile, with the majority of mice not suffering severe pancreatitis or hysterosalphingitis in the first 5 months of life. The mean lifespan of TNF-/- pfp-/- gld mice was 217 +/- 79 days compared with 69 +/- 10 days for TNF+/+ pfp-/- gld mice and the majority of moribund TNF-/- pfp-/- gld mice appeared to die as a result of severe pancreatitis, suggesting that loss of TNF was not completely protective. At 8 weeks of age, characteristics associated with the gld phenotype, such as expansion of B220+ CD4- CD8- T cells, lymphadenopathy and hypergammaglobulinemia were comparable between TNF+/+ pfp-/- gld and TNF-/- pfp-/- gld mice, although the lymphoid organs of TNF+/+ pfp-/- gld mice contained greater numbers of B220+ CD4- CD8- T cells, macrophages and T cells. We conclude that TNF is necessary for the full manifestation of immune dysregulation caused by pfp/FasL-deficiency, in particular in the early and overwhelming tissue infiltration and destruction caused by inflammatory cells.

[Functional activity of phagocytizing cells of the female reproductive tract in inflammation of the upper part of genitalia]. / Funktsional'naia aktivnost' fagotsitiruiushchikh kletok reproduktivnogo trakta zhenshchin pri vospalenii verkhnego otdela genitalii.

66 women of reproductive age with different course of the inflammatory process in the upper section of the reproductive tract (endometritis and salpingo-oophoritis) were examined. The cell composition, viability and functional activity of the phagocytizing cells of cervical and endometrial secretions, as well as peritoneal exudate, were studied. The study revealed that these characteristics of the phagocytizing cells of the reproductive tract in women with the inflammatory process differed from similar characteristics in healthy women. Different changes in the functional activity of neutrophils and macrophages in the biological fluids under study in different course of the inflammatory process were detected.

Perforin/Fas-ligand double deficiency is associated with macrophage expansion and severe pancreatitis.

We report that perforin/Fas-ligand double-deficient mice die early of severe pancreatitis. Female mice, in addition, are infertile and suffer from hysterosalpingitis. Tissue destruction is accompanied by infiltration with Mac-1 (CD11b)-positive monocytes/macrophages, Mac-1-positive T cells, and expansion of CD8+ T cells. In vivo inactivation of monocytes/macrophages by carrageenan reverses disease progression and restores fertility of female mice. Perforin/Fas-ligand double-deficient CD4+ or CD8+ CTL are unable to lyse cognate-activated macrophages, and therefore are unable to mediate negative feedback regulation by lysis of APCs, thereby preventing further T cell activation. These studies demonstrate a novel role for perforin in homeostatic regulation of the immune response.

[Local humoral immunity in women with combined forms of infertility]. / Mistsevyi humoral'nyi imunitet u zhinok z poiednanymy formamy neplidnosti.

An evaluation was done of the immunity status in 100 women presenting with infertility of associated genesis. An immunologic investigation into blood serum, follicular fluid (FF) and peritoneal fluid (PF) was carried out at day 14-16 and 18-23 of the menstrual cycle. The functional status of spermatozoa was characterized with the aid of the index of their survival in FF and PF. Local humoral immunity of FF and PF differs from the indices in blood serum, which fact was manifested by a striking reduction of Igs G, A, M. Sperm-immobilizing antibodies (SIAB) have been identified in blood serum, FF, and PF in 14, 4%, 22.4%, and 25% of women. SIAB production is associated with inadequacy of local immunity, such as low levels of SIgA, IgA, and lysozyme. Occurrence of SIAB in PF and FF worsens general motility of spermatozoa by agglutination of the latter, which fact undoubtedly affects conception.

Serological responses of patients with ectopic pregnancy to epitopes of the Chlamydia trachomatis 60 kDa heat shock protein.

Clinical and histopathological correlations of immunoreactivity to Chlamydia trachomatis and to epitopes of the C. trachomatis 60 kDa heat shock protein (hsp60) among women with ectopic pregnancy were evaluated in a case-control study. Serological responses to 13 synthetic peptides corresponding to major epitopes of the chlamydial hsp60 were determined in 67 women treated for ectopic pregnancy and 45 women with uncomplicated pregnancy in utero. Plasma cell salpingitis was detected in 29 (43.3%) of the ectopic patients. Its presence correlated with antibodies to two hsp60 epitopes, encompassing amino acids 260-271 and 411-422 (P = 0.02). Antibodies to these two epitopes, along with five other epitopes, also correlated with peritubal adhesion formation in ectopic pregnant patients (P < 0.01). Antibodies to epitopes 260-271 and 188-199 also correlated with a history of pelvic inflammatory disease (PID; P = 0.05). Patients with ectopic pregnancy were also more likely than their intrauterine pregnant controls to have present anti-chlamydial immunoglobulin G (P < 0.005). Women positive for both C. trachomatis and hsp60 epitope antibodies had an increased prevalence over controls of salpingitis, pelvic adhesions or history of PID (P < 0.05). In contrast, patients who were positive for only C. trachomatis antibodies or only hsp60 epitope antibodies did not differ from antibody-negative patients in each of these categories.

Antibodies to the chlamydial 60 kd heat-shock protein are associated with laparoscopically confirmed perihepatitis.

OBJECTIVE: Our purpose was to examine clinical, microbiologic, serologic, and laparoscopic findings associated with perihepatitis. STUDY DESIGN: In a prospective study of 157 women with a clinical diagnosis of pelvic inflammatory disease, 27 women with laparoscopically confirmed perihepatitis and salpingitis were compared with 46 patients with salpingitis alone. RESULTS: Both current use or a history of ever using oral contraceptives was negatively associated with perihepatitis (p = 0.05 and p = 0.008, respectively). Moderate-to-severe pelvic adhesions were present at laparoscopy significantly more often in the perihepatitis-salpingitis group (70%) than in the salpingitis alone group (35%, p = 0.003). Antibody to the chlamydial 60 kd heat-shock protein at > or =0.5 optical density was detected in 67% of the perihepatitis-salpingitis group and in 28% of the salpingitis alone group (p = 0.005), and the median titer was significantly higher in the former group (p = 0.02). CONCLUSION: Compared with women with salpingitis alone, patients with perihepatitis-salpingitis do not have distinctive clinical or microbiologic findings but do manifest a higher prevalence of moderate-to-severe pelvic adhesions and both a higher prevalence and higher titers of antibody to the chlamydial heat-shock protein-60.

Humoral immune response to plasmid protein pgp3 in patients with Chlamydia trachomatis infection.

We identified, by two-dimensional electrophoretic analysis and microsequencing, a protein of Chlamydia trachomatis elementary bodies which corresponds to the polypeptide (pgp3) encoded by open reading frame 3 (ORF3). Amino acid analysis showed that the first residue (Gly) found in the native protein is the one encoded by the second ORF3 codon, implying a typical bacterial removal of the first Met residue. Relatively large amounts of recombinant pgp3 (r-pgp3) in a stable, water-soluble form were obtained by overexpressing ORF3 in Escherichia coli and purifying the product from periplasmic extracts under nondenaturing conditions. These r-pgp3 preparations allowed specific detection of anti-pgp3 antibodies by enzyme-linked immunosorbent assay. Analysis of a group of 170 sera from healthy blood donors and from patients who were seropositive or -negative for C. trachomatis and Chlamydia pneumoniae showed that an immune response to pgp3 occurs in the majority (ca. 81%) of patients with sexually transmitted diseases who are seropositive for C. trachomatis and generally correlates with the response to cell surface antigens. No reaction between r-pgp3 and C. pneumoniae-positive sera was detected.

In situ characterization of leukocytes in the fallopian tube in women with or without an intrauterine contraceptive device.

Histological evaluation of sections from the human fallopian tube revealed an inflammatory reaction in 21 of 31 women using an intrauterine contraceptive device (IUCD) and in four of 29 controls (non-IUCD users). The inflammatory cells were mainly localized at the epithelium-lamina propria interface and at the center of the mucosal folds. The immunohistochemical study revealed leukocytes (CD45+), T lymphocytes (CD3+), T helper cells (CD4+), T suppressor/cytotoxic cells (CD8+), B lymphocytes (CD22+, CD19+), granulocytes, monocytes and null cells (CD11b+) mainly localized at the lamina propria in both groups. T lymphocytes were the predominant cell type, and the ratio between T helper and T suppressor/cytotoxic cells was fairly close to one both in IUCD-users and controls. B lymphocytes were the least frequent cell type identified. In IUCD users, the numbers of the different leukocytes were increased. In both groups, IgA-, IgG- and IgM-positive cells were demonstrated and were predominantly located at the lamina propria of the mucosal folds. The IgA-positive cells dominated in both groups, whereas IgG- and IgM-positive cells were less frequent. Cell positive for IgA, IgG or IgM were significantly increased in number in the IUCD users. The data confirm the presence of an immune system in the normal human fallopian tube and indicate that the IUCD can induce a prominent recruitment of inflammatory cells, with a tubal inflammation as the result. The IUCD may disturb the immunological function of the fallopian tube and its rôle in fertilization.

PIP: In Norway, a 1.5 cm segment of the isthmic portion of the fallopian tube was removed for immunohistochemical and histological analyses from 60 healthy nonpregnant women. Researchers compared data on the 31 women who had used a copper IUD for 2-10 years with data on the remaining 29 women who did not use any IUD for the last year to examine the localization, number, and subtypes of leukocytes in the tube. An inflammatory reaction was present in 68% of the IUD users compared to only 14% of controls (p = 0.005). A chronic inflammatory reaction was the predominant type in both groups (52% for IUD users and 8% in controls). The inflammatory cells were largely found in the epithelium-lamina propria interface and in the center of the mucosal folds. In both groups, leukocytes (CD45+), T lymphocytes (CD3+), T helper cells (CD4+), T suppressor/cytotoxic cells (CD8+), B lymphocytes (CD22+, CD19+), granulocytes, monocytes, and null cells (CD11b+) were chiefly located at the lamina propria. CD3+ cells were the primary cell type in both groups. The ratio of CD4+ and CD8+ was close to one in both groups. CD22+ and CD19+ were relatively rare in both groups. The number of the different leukocytes was significantly greater among IUD users than the controls (e.g., total leukocytes, 1127 vs. 609; p 0.0001). The lamina propria of the mucosal folds housed most IgA, IgG, and IgM positive cells. IgA positive cells were the most frequent cells in both groups. The number of cells positive for IgA, IgG, or IgM were significantly higher in the IUD user group than the control group (e.g., total Ig in endosalpinx, 188 vs. 50; p 0.0001). These findings show that the IUD can incite inflammatory cells, resulting in inflammation of the fallopian tube. Thus, the IUD may interfere with the immunological function of the tube and the tube's role in fertilization.

Analysis of the immune response in mice following intrauterine infection with the Chlamydia trachomatis mouse pneumonitis biovar.

A Swiss Webster white mouse model of salpingitis was used to characterize the immune response following an intrauterine infection with the Chlamydia trachomatis mouse pneumonitis biovar. Western blot (immunoblot) analyses of the serum samples showed that the immunodominant bands corresponded to molecular masses of 72, 60, 42, and 28 kDa and to the lipopolysaccharide. Antibodies to the 60-kDa heat shock protein and to the 60-kDa cysteine-rich protein were detected at 2 and 3 weeks postinfection, respectively. Neutralization was observed in an in vitro assay with serum samples as early as the 3rd day postinfection and remained high for the 7 weeks of observation. The mice were mated in the 7th week following infection. Of the infected experimental mice, 71.4% were found to be either unilaterally or bilaterally infertile, whereas only 27.4% of the noninfected control mice were found to be infertile.

In vivo tumor necrosis factor production in women with salpingitis.

Immune mediated mechanisms might contribute to damage of the fallopian tube in instances of salpingitis. Using a filter paper technique to obtain samples during the surgical procedure, we examined fluids from the reproductive tract organs of seven women with salpingitis and five controls, for evidence of tumor necrosis factor (TNF). TNF, produced principally by macrophages, is a substantial mediator of inflammatory responses. In three women culture-positive for Chlamydia trachomatis, TNF was identified only in those fallopian tubes with visual evidence of disease. Fluids obtained from morphologically normal tubes, as well as from the ovaries and uterus, were negative. In three women with negative fallopian tube cultures but visual evidence of salpingitis, TNF was also identified in fluids from damaged, but not from normal, tubes. Ovarian and uterine fluids of the women were also TNF positive. The last patient, also culture-negative, had TNF only in one affected tube. All five patients in the control group had negative findings at all genital tract sites. Only one patient had TNF in her serum. Thus, localized cell-mediated immune system activation, identified by TNF production, appears to be a typical component of salpingitis.

[Cultural and serologic Chlamydia detection in diagnosis of sterility and increased risk of infection]. / Kultureller und serologischer Chlamydien-Nachweis in der Sterilitätsdiagnostik und bei erhöhtem Infektionsrisiko.

364 women were studied for the possible role of chlamydia trachomatis infection as a factor related to infertility. C. trachomatis was isolated from the cervix in 20% of our high-risk STD patients and in a much lower proportion in patients with tubal (5%) or unexplained (4%) infertility. IgG-antibodies to C. trachomatis were found in 75% of patients with tubal infertility, in 44% of women without gynaecological diseases and in 87% of the high-risk STD patients. By contrast, the difference was significant in the IgA-antibody tests. 67% of the women with tubal infertility and only 25% of the women with nontubal infertility were positive. From these data it appears, that antecedent infections with C. trachomatis, as measured by antibody prevalence, are an important factor in infertility of tubal origin.

High hepatitis B carrier rate among non-Oriental patients with acute salpingitis.

Hepatitis B surface antigen screening was performed on non-Oriental patients with the diagnosis of acute salpingitis. Six of 59 (10.17%) had hepatitis B surface antigenemia. We therefore recommend hepatitis B screening for all patients with salpingitis. The detection of covert hepatitis B helps to protect hospital staff and provides important information for the patient. Further, the administration of hepatitis B vaccine may be appropriate for those patients with acute salpingitis who are found to be hepatitis B surface antigen-negative.

Serum CA 125 in acute pelvic inflammatory disease.

Serum levels of CA 125 and other selected tumour markers were measured in 31 patients with proven pelvic inflammatory disease (PID). Ten (32%) of the patients had elevated CA 125, one (4%) had elevated CEA, and none had elevated CA 15-3, AFP or beta 2-microglobulin. Compared to patients with normal CA 125, patients with elevated CA 125 were older, more often users of intrauterine contraceptive devices, had longer duration of symptoms, higher erythrocyte sedimentation rates, and more often had an adnexal mass on pelvic examination. There was a correlation between CA 125 levels and the severity of adnexal inflammation as defined by laparoscopy. Isolation of specific micro-organisms from the upper genital tract was not associated with elevated CA 125. In most women serum levels of CA 125 decreased during treatment. PID should be considered as a major cause of positive CA 125 findings among young women.

Detection of specific IgG and IgA antibodies to Chlamydia trachomatis in women with salpingitis confirmed by laparoscopy.

The sera from 12 consecutive symptomatic women with laparoscopy-confirmed salpingitis were screened for the presence of specific IgG and IgA antibodies to Chlamydia trachomatis by a single antigen (L-2) immunoperoxidase assay. All women were found to have IgG and IgA antibodies to C trachomatis. Six women had positive endocervical cultures for C trachomatis, and one of these had positive cultures from the conjunctiva and fallopian tubes. Serum chlamydial IgA antibodies may serve as markers for active infection with C trachomatis regardless of whether organisms can be identified by culture or direct fluorescent antibody techniques from endocervical or fallopian tube samples.