1. Concentration Levels of IL -10 and TNF-α Cytokines in Patients with HPV DNA Positive and Negative Cervical Lesions*
Katan Sabir Ali *, Husham Y. M. Ali Bayazed**, Jaladet M. S. Jubrael***
*College of Medicine, Hawler Medical University,
** Consultant / Scientific Research Centre / University of Zakho, *** Molecular Biology Central Laboratory, Duhok university,
Ministry of Higher Education & Scientific Research, Kurdistan Regional Government- Iraq

2. Introduction
More than 100 different types of HPV have now been identified by molecular technique and those infecting the genital tract are divided into low risk (LR) and high risk (HR) types. The LR types induces only benign genital warts and include HPV 6 and 11, while the HR types such as HPV 16 and 18 types are associated with the development of cervical cancers. The cellular immunity plays a important role in the early phase of infection and it is the only efficient response against non-lytic HPVs and is involved in the immune response to neoplasia. The immune response is mediated through the release of different mediators, which can influence the synthesis of different immunoregulating cytokine networks.

3. Materials and Methods
The study was carried out on 80 females, with different cervical lesions, and 20 healthy individuals as control group with age range of years. The enrolled cases were attendees of the Maternity Hospital in Hawler city, Kurdistan Regional Government, Iraq . All patients were interviewed, underwent full clinical examinations by a specialist gynecologist, and some relevant laboratory investigations were performed in addition to Pap smears. The cervical smears were obtained and cytologically classified by a specialist pathologist into five categories: 20 with atypical squamous cells of undetermined significance (ASCUS), 30 with LSIL (CIN I), 16 with HSIL (CIN II and CIN III), 14 with SCCC (squamous cells cervical carcinoma), and 20 with normal cervical cytology (control group). Molecular technique was performed utilizing PCR amplification kits (Sacace Biotechnologies ,Caserta, Italy Catalog No.V-26-50F) and PCR amplification kit, HPV High Risk Typing, (Sacace Biotechnologies ,Caserta, Italy Catalog No.V-25-50F) according to manufacturer’s instructions.

4. Cervical secretions and blood samples were obtained from all participants. As much as possible cervical secretions were collected prior to the Pap smear by inserting 2 ml of sterile phosphate-buffered saline (PBS) using a sterile disposable plastic Pasteur pipette. The cervical fluid were processed immediately in the laboratory by centrifugation at 3500rpm for 10 minutes and the supernatants stored at -20 °C until analysis at the laboratory center of Nanakali Hospital. The obtained cervical secretions with blood samples from the studied cases (HPV positive and negative cases and control group) were tested for estimation of IL-10 and TNF-α concentration levels, utilizing ELISA kits (IMMUNOTECH, ABEKMAN CULTURE COMPANY IMMUNOTECH.S AS Marseille Cedecx 9 France).

5. Results
The detected levels of IL-10 (mean ± SE) concentration in cervical secretions of patients with HPV DNA positive and negative states and control group were ± pg/ml, ± 2.84 pg/ml and 8.27 ± 0.59 pg/ml respectively with significant differences (p<0.05), while levels of TNF-α in cervical secretion of the studied groups were ± 3.49 pg/ml, 9.90 ± 0.73 pg/ml, and 7.90 ± 0.87 pg/ml respectively with non significant differences. The detected levels of IL-10 in cervical secretions of HPV DNA positive cases (88.73 ± pg/ml) were significantly higher than in the sera (13.69 ± 2.41 pg/ml) (p<0.05), while the levels of TNF-α in their cervical secretions (12.18 ± 3.49 pg/ml) was slightly raised than in their sera ( ± 3.14 pg/ml) with non significance differences.

6. Discussion & Conclusion
A raised levels of both IL-10 and TNF-α in secretions of HPV D NA positive women with different cervical lesions were detected. However, the observed higher levels of IL-10 than TNF-α indicate down-modulation of tumor-specific immune response to HPV infected lesions via significant raised concentrations of the first cytokine than the second one. Therefore, this phenomenon seems to provide a tumor progressive microenvironment by the immunosuppressant properties of IL-10 with minimal antitumor activity of TNF-α.

7. These findings highlight a local immune response to virally infected lesions with a shift of cytokine profile production to immunosuppressive status in lesions with different stages, but more accentuated in progressive lesions. Similar role of HPV infected cervical lesions with associated lesions dysplasia and viral immune modulation and evasion of host immune response were documented by others . Moreover, IL-10 was found to be is a pleiotropic cytokine and proved to have immunosuppressive effects on the cervical epithelium with increase in its expression levels that hasten lesion progression from normal cervical epithelium to low- grade lesions and consequently to high-grade lesions and cancer stage .

8. Author Affiliation
Prof Dr Husham Bayazed MBChB, MSc, PhD - Consultant of Immunology & Infectious Diseases Scientific Research Center/ University of Zakho - Fellowship of International Society of Chemotherapy, Infection, Cancer (Canada) - Immunology Advisory Board Member (EUROMDnet) (Belgium) - Membership of World Stroke Organization (Geneva) - Membership of Metabolomics Society (USA) - Membership of American Association of Science & Technology - PubMed Medline: Ali HY