Subbi Mathur

Cyclic variations in white cell subpopulations in the human menstrual cycle: Correlations with progesterone and estradiol

Cyclic variations in white cell subpopulations were studied in serial blood samples from 18 female volunteers (14 ovulatory and 4 nonovulatory cycles) and 2 males. Total white blood cells (WBC), lymphocytes, total and active T cells (TEt, TEa), monocytes, and granulocytes were counted, and levels of estradiol (E2), progesterone, and luteinizing hormone (LH) were measured. In the ovulatory cycles, lymphocyte counts at midcycle (Day 0) reached a minimum, coinciding with the maximum level of E2 peak (35 ng%). In daily samples, the minimum lymphocyte counts coincided with the preovulatory E2 surge (P < 0.01). Similar but less significant negative correlations (P < 0.05) were found between E2, WBC, and TEt. In contrast, TEa did not show cyclic variations. Monocyte and granulocyte counts were significantly higher in the luteal than in the follicular phase (P < 0.05); their pattern followed closely that of progesterone (P < 0.05) but not of E2. Daily evaluations of hematocrit, hemoglobin content, red blood cell count, and mean corpuscular volume failed to show cyclic variations. For the females with nonovulatory cycles and for the males studied (with sex steroid profiles corresponding to the follicular phase), no cyclic variations were found in white cell subpopulations.

Autoimmunity in endometriosis: relevance to infertility.

PROBLEM: Endometriosis is a complicated multi‐factorial disease. Genetic and immunologic factors play a key role in its pathophysiology. We were the first to describe endometrial and ovarian autoimmunity in women with endometriosis in 1981.

Clinical significance of sperm antibodies in infertility

Sperm antibody (AB) titers, determined by passive hemagglutination and cytotoxicity assays, were found to be elevated in 62 males and 46 females of 103 couples with primary infertility; 15 males and 12 females of 25 couples with secondary infertility; 10 males and 8 females of 18 couples with histories of repeated abortion; 21 males and 17 females of 25 couples in which the husband had a history of prostatitis; and 29 males and 17 females of 38 couples in which the husband had oligospermia. Of the couples in which one or both partners had elevated sperm AB titers, only 4 achieved pregnancy: 3 from the group with secondary infertility, all of which ended in spontaneous abortions, and 1 in which the husband was oligospermic. This suggests an etiologic role of sperm immunity in infertility. Immunosuppressive treatment of autoimmune males with prednisone (15 mg/day for 3 weeks to 6 months) resulted in significant decreases in AB titers. Pregnancies were achieved by 9 of 25 couples after treatment (36%). The observed increase in pregnancy rate in the prednisone-treated versus untreated groups of couples with elevated sperm AB titers was significant (P less than 0.02).

Endometrial antigens involved in the autoimmunity of endometriosis.

Serum and peritoneal fluid from five fertile women without endometriosis and serum (n = 23) and peritoneal fluid (n = 12) from infertile women with endometriosis were tested for the presence of antibodies against endometrial tissue antigens by a Western blot analysis. Antigens with molecular weights (MW) of 19, 31, 38, and 42 kd reacted with antibodies in the serum and peritoneal fluid from both fertile and infertile women. Antibodies in 20 of 23 (87%) sera and all 12 (100%) peritoneal fluid samples from endometriosis patients reacted against endometrial antigens with molecular weights (MW) of 26 kd and/or 34 kd. Serum from 10 patients (43%) and peritoneal fluid from 6 patients (50%) also had antibodies to an endometrial antigen with MW of 21.5 kd. Reactivity to other endometrial antigens with MW 16, 24, 48, and 75 kd was also noted in patients with endometriosis. Antibodies in the serum and peritoneal fluid from fertile women failed to react against these antigens. It is concluded that the humoral and local endometrial autoimmunity detected in patients with endometriosis is primarily directed against antigens with MW of 26 and 34 kd.

An endometrial antibody assay in the clinical diagnosis and management of endometriosis

Coded serum samples from 11 normal fertile men and 17 fertile women without endometriosis (control groups) and 41 women with endometriosis were tested blindly for the presence of endometrial antibodies by use of a passive hemagglutination assay. Endometrial antibodies were either absent or present in low baseline titers in the serum samples from the control group. In contrast, 17 of the 23 (74%) patients with untreated endometriosis had elevated titers of endometrial antibodies in their serum. Of the 18 patients treated with danazol, endometrial antibodies were absent in 7 women who showed a good response at repeat laparoscopy, whereas 4 of 5 women with a poor response had significantly positive titers of antibodies. Six patients treated with danazol did not have repeat laparoscopy, but were found to have endometrial antibody titers in the baseline control range. Endometrial antibody titers in women with a good response to danazol were significantly lower than those in women with untreated endometriosis or with a poor response to danazol (P = 0.003). No correlation was observed between the antibody titers and the stage of endometriosis. The results suggest that the assay for serum endometrial antibodies may prove to be a clinically useful, noninvasive aid in the diagnosis of endometriosis. Sequential determination of endometrial antibody titers may be helpful in assessing the efficacy of pharmacologic therapy for endometriosis and evaluating the cases of patients with possible recurrence of the disease.

Cross-reactivity of sperm and T lymphocyte antigens.

ABSTRACT: Evidence is presented for cross‐reactivity between antigens on human sperm and T lymphocytes. In 25 infertile couples in which both the males and females had significant antisperm immunity, antibody (Ab) titers to thymocytes (mean ± S.E.M. 159 ±4 and 72 ± 14, respectively, in males and females), T cell lines CCRF‐CEM (69±5 and 48±8) and HSB‐2 (56±15) and 41±8), suppressorenriched (TG) cells (26±6 and 66±28) and helper‐enriched (Tg—) cells (26±4 and 46±14) were significantly elevated, as compared wth Ab titers in 45 normal males and 45 normal females without antisperm immunity. Antibody titers to adult B cells, B cell line RAJI, and granulocytes were similar in the two groups. Antisperm Ab titers in sera, sperm extracts, and seminal plasma of the infertile subjects were significantly reduced after absorption with sperm, thymocytes, or T cell line CCRF‐CEM but not with the B cell line RAJI. Antithymocyte Ab titers in the sera were significantly reduced (p < 0.001) after absorption with thymocytes, CCRF‐CEM, or sperm, but not RAJI. Lymphocytes from the infertile patients, when stimulated with pokeweed mitogen in vitro, produced antisperm and anti‐T‐lymphocyte antibodies at significantly higher titers than normal controls.

Sperm motility on postcoital testing correlates with male autoimmunity to sperm

To assess whether or not immunologic factors in husbands, wives, or both, influence the motility of sperm in the female reproductive tract, hemagglutination and cytotoxicity sperm antibody (Ab) assays and postcoital tests (PCTs) were performed in 293 infertile couples. More couples without male autoimmunity to sperm (64% of 66; P less than 0.001) had greater than or equal to 10 motile sperm per high power field (adequate PCT), as compared with 26% of 122 couples with untreated male autoimmunity, 19% of 77 couples with corticosteroid-treated male autoimmunity without a pregnancy, and 36% of 28 couples with successfully treated male autoimmunity to sperm. Good correlation was obtained among pregnancy achievement, lack of sperm antibodies, and adequate sperm motility in the PCT (P less than 0.0001). Sperm motility in the PCT correlated positively with sperm motility in the semen and inversely with cytotoxic sperm Ab in the serum and seminal plasma of men and women and hemagglutinating sperm Ab in the cervical mucus samples. Sperm motility in the PCT has a predictive value of 72% for male autoimmunity and 57% for female isoimmunity to sperm in the presence of normal cervical mucus and in the absence of infections.

Cervical epidermal growth factor-receptor (EGF-R) and serum insulin-like growth factor II (IGF-II) levels are potential markers for cervical cancer.

PROBLEM: To ascertain if cervical epithelial epidermal growth factor receptor (EGF‐R) and serum insulin‐like growth factor II (IGF‐II) levels are potential markers for cervical cancer.
 METHOD OF STUDY: We tested cervical biopsies obtained from 18 controls, 3 women with cervical intraepithelial neoplasia (CIN) I, 17 women with CIN II and III, and 12 women with cervical cancer for EGF‐R using a quantitative immunofluorescent antibody assay. We measured serum IGF‐II levels using an enzyme‐linked immunosorbent assay in 20 controls, 26 CIN patients, 12 with cervical cancer before therapy, 5 with cervical cancer for <1 year, and 9 others ≥1 year after therapy.
 RESULTS: The levels of cervical EGF‐R in women with CIN and cervical cancer were significantly higher (P<0.05 for CIN I; P<0.001 for patients with CIN II and III or cervical cancer) than in controls. Women with cervical cancer (P<0.001 vs. controls) or advanced CIN (P=0.03) had elevated levels of serum IGF‐II, while the women with CIN I had levels similar to controls. Women with cervical cancer in the post‐therapy period had significantly lower serum IGF‐II levels than the women with cervical cancer before therapy (P<0.001).
 CONCLUSION: Cervical epithelial EGF‐R and serum IGF‐II levels may be used for the diagnosis and prognosis of cervical cancer.

Human Papilloma Virus (HPV)-E6/E7 and Epidermal Growth Factor Receptor (EGF-R) Protein Levels in Cervical Cancer and Cervical Intraepithelial Neoplasia (CIN)

BACKGROUND: About 90% of cervical cancers and advanced cervical intraepithelial neoplasia (CIN II/III) are squamous epithelial cells with mRNA for human papillomavirus (HPV)16 and 18 and up‐regulated epidermal growth factor receptor (EGF‐R). Since presence of proteins rather than mRNA may be truly indicative of active infection or disease progression, establishing reliable methods for quantifying these proteins in cervical biopsies is important.
 METHOD: We have established an objective semi‐quantitative immunofluorescent antibody assay to reliably assess the levels of HPV‐E6/E7 and EGF‐R proteins in the cervical biopsies from 12 normal women, five women with CIN I, 15 with CIN II/III and ten with cervical cancer.
 RESULTS: HPV‐E6/E7 and EGF‐R, when present, were specific to para‐basal, basal and squamous epithelial cells (negative in stromal cells). Nine of ten women with cervical cancer and 15 (14 CIN II/III; 1 CIN I) of 20 women with CIN were positive for HPV‐E6/E7. All 12 controls were HPV‐negative. The controls and six women with CIN (four with CIN I) negative for HPV had low levels of EGF‐R. The only exception was one woman with cervical cancer negative for HPV, with high levels of EGF‐R. Levels of HPV‐E6/E7 and EGF‐R were significantly higher (P<0.001 vs. controls) in women with advanced CIN II and III (P<0.05 vs. controls in CIN I) and cervical cancer. The HPV‐E6/E7 and EGF‐R levels correlated significantly (r=18.98; P<0.001, by linear regression analysis).
 CONCLUSION: We have established a highly specific and sensitive semi‐quantitative immunofluorescent antibody assay for measuring levels of HPV‐E6/E7 proteins and EGF‐R in archival cervical biopsies. Our data suggest an association between HPV‐E6/E7 and EGF‐R.

Levels of Transferrin and Alpha 2-HS Glycoprotein in Women with and without Endometriosis

The study objective was to test the hypothesis that elevated levels of transferrin and alpha 2-HS glycoprotein occur in the peritoneal environment of patients with endometriosis that may lead to the observed autoimmunity to these proteins. We set up a double sandwich enzyme-linked immunosorbent assay (ELISA) for measuring levels of transferrin and alpha 2-HS glycoprotein in the serum and peritoneal fluid samples from women with (n = 24-60) and without endometriosis (n = 35-49). Serum and peritoneal fluid levels of alpha 2-HS glycoprotein and peritoneal fluid levels of transferrin were significantly elevated in patients with endometriosis, in contrast to the controls. Serum levels of transferrin in patients, however, were significantly less in the patients than in controls. We conclude that transferrin and alpha 2-HS glycoprotein are present at high concentrations in the peritoneal fluids of patients with endometriosis. This may play a significant role in the autoimmune pathophysiology of endometriosis.