Usefulness of Positron Emission Tomography for genital Chlamydia infection assessment in the murine model.

Usefulness of Positron Emission Tomography for genital Chlamydia infection assessment in the murine model. Marangoni A.1, Nanni C. 2, Quarta C. 2, Foschi C. 1, Russo I. 1, Nardini P. 1, D’Errico A. 3, Rosini F. 3, Aldini R.4, Cevenini R.1 1Section of Microbiology-DESOS, 3Med. Interna, Dell’invecchiamento e Malattie nefrologiche, 4SMETEC Dpt., University of Bologna. 2Nuclear Medicine, Azienda Ospedaliero-Universitaria di Bologna, Policlinico S.Orsola-Malpighi, Bologna, Italy. OBJECTIVES Untreated Chlamydia trachomatis infection can wreak havoc on the reproductive organs profoundly affecting fertility in women. Taken together, the high rate of asymptomatic infections and the severity of the infection related pathology indicate that control of chlamydial infections would require the development of new diagnostic non-invasive techniques for genital infection. The aim of this study was to explore the feasibility of 11C-choline Positron Emission Tomography (PET) in the assessment of the degree of inflammation in a C. muridarum genital infection mouse model. METHODS Model of infection. All the experiments were approved by the Ethical Committee of the University of Bologna. Animals used were 43 female Balb/c mice, 6-8 weeks old. All animals received 2.5 mg of medroxyprogesterone acetate i. m. 9 and 2 days prior the infection. Twenty-one mice were infected by placing 15 µl of sucrose-phosphate-glutamic acid (SPG) buffer containing 106 inclusion forming units (IFUs) of C. muridarum into the vaginal vault. Twelve animals were treated with 15 µl of SPG containing heat-inactivated 106 IFUs of C. muridarum. As controls of inflammation, 7 animals were challenged with 15 µl of SPG. The Experimental design. Twenty-one infected animals were randomly allotted into two groups. Group A: 9 animals underwent a 11C-Choline PET at day 5, 10 and 20 post-infection. Group B: 12 animals were sacrificed at 5, 10, 15, 20 days for culture and histological analysis (3 animals for each time point). The 12 mice treated with inactivated chlamydiae were divided in two groups, as well. Group C: 4 animals underwent a 11C-Choline PET at day 5, 10 and 20 post-inoculation. Group D: 8 animals were sacrificed at 5, 10, 15, 20 days for culture and histological analysis (2 animals for each time point). Five control animals underwent 11C-Choline PET at the same days of the infected animals. Similarly, the remaining two control animals were sacrificed for histological analysis, one at 10 days and the other one at 20 days post-inoculation. PET. 11C-Choline PET was carried out as follows: each animal was anaesthetized with gas anesthesia (Sevofluorane 3–5% and oxygen 1 l/min) and was injected with approximately 20 MBq of 11C-Choline, in a volume of 0.1 ml. The residual dose was measured to verify the effective dose injected. The animal was subsequently placed on the scanner bed in the prone position, after an uptake time of 10 minutes. Images were acquired with a small animal PET tomograph for a total acquisition time of 15 min. 11C-Choline PET images were reconstructed iteratively (OSEM 2D) and read in three planes (axial, sagittal and coronal). Semi-quantitative analysis was carried out using the parameter SUV (standardized uptake value) representing mean radioactive counts per gram of tissue, divided by injected dose of radioactivity per gram of animal weight. The target region of interest (ROI) was placed on the pelvis at the level of the cervical-vaginal region. Histology. The specimens obtained from sacrificed animals were preserved in 10% neutral buffered formalin for about 48 h and put in embedding cassettes, then processed in automatic tissue processor. After dehydration they were infiltrated with molten paraffin wax. Four-µm thick sections were cut with microtome, stained with haematoxylin-eosin and mounted on glass microscope slides. The uterine horns were scored with respect to inflammation using the following criteria: 0: No inflammation. 1...