INTRODUCTION:

Regardless of surgical or non-surgical intervention, anterior cruciate ligament (ACL) rupture is associated with the development of post-traumatic osteoarthritis (PTOA) within 5-10 years of the incident injury^1-3. As such, recent efforts have focused on identifying risk factors associated with ACL-injury, with the goal of reducing its overall incidence. Previous work has identified both intrinsic and extrinsic factors that contribute to ACL-injury risk, and multiple studies have established that female ACL-injury risk is more than double that of their male counterparts^4-7. Sex based disparities in ACL-injury risk may be explained by cyclic variations in hormones, as previous studies that have identified that female ACL-injury risk is increased (along with anterior knee laxity) during the pre-ovulatory phase of the menstrual cycle^{5, 6}. These findings suggest that sex hormones may directly affect the structure and biomechanical integrity of the ACL. Ultra-short echo (UTE) MRI sequences can be utilized to capture the rapid transverse relaxation times associated with ligaments and allow for quantitative evaluation of tissue microstructure with T2* mapping^8-12. Recently, UTE-MRI and bi-component T2* analyses have been used to evaluate both the short T2* (associated with bound water) and long T2* components (associated with free water) within tissues and assess their relationship to mechanical properties^{8, 9}. The objective of this study was to determine if significant changes in ACL T2* metrics exist over the course of the female menstrual cycle. We hypothesized that normally ovulating pre-menopausal females would exhibit significant changes in ACL T2* metrics over the course of a menstrual cycle, while no such differences would exist within anovulatory control subjects.

METHODS:

This was an IRB approved pilot study that included 23 females with no history of knee injury. Nine pre-menopausal females with normal menstrual cycles and no history of hormonal contraceptive use (>1 year) were included as ovulatory case subjects. Subjects within the anovulatory control group included 6 post-menopausal females, and 8 pre-menopausal subjects taking oral contraceptives with normal menses. All subjects participated in 4 study visits evenly spaced over the course of 1-month (1 per week). Study visit #1 for all pre-menopausal females coincided with onset of menses (within 24hrs). Subjects were provided with commercially available ovulation predictor kits (ClearBlue Digital Ovulation Tests [Accuracy 99%]) to determine the date of ovulation in case subjects, and to confirm anovulatory status in control subjects. MRI Acquisition: At each study visit, bilateral MRI examinations were obtained on a 3-Tesla clinical scanner (GE Healthcare) using an 8-channel phased array knee coil (Invivo). Three-dimensional, double obliqued (to the ACL in axial and sagittal planes) coronal UTE sequences were acquired for the evaluation of T2* metrics (Voxel: 0.50x0.50x1.5mm³, TEs: 11 echoes between 0.03-25ms, TR: 166ms, RBW: ±83.3kHz, Flip-Angle: 16^o). Imaging Analysis: Bi-exponential fits of SI to corresponding echo time were used to calculate ACL T2* metrics: SI(TE) = A(-TE/T2*S) + B(-TE/T2*L)+noise, where T2*S and T2*L are respective short and long T2* components, A and B are corresponding short and long apparent proton densities, and percentage short (PS) ¹² is calculated as A/(A+B). Statistical Analysis: paired and unpaired t-tests were utilized for the evaluation of within and between group differences across all study visits.

RESULTS:

Significant changes in ACL T2* metrics over time were found within ovulatory case subjects only. Specifically, ovulatory case subjects exhibited significantly decreased mean and median T2*S and median PS metrics within the pre-ovulatory phase compared to the post-ovulatory phase (T2*S mean difference: = -0.85ms; T2*S median difference = -1.54ms; PS median difference = -2.47, p ≤ 0. 05). Anovulatory control subjects exhibited no significant changes over time for any T2* metric. (Figure 1)

DISCUSSION:

ACLs of normally ovulating, pre-menopausal case subjects displayed significant changes in mean and median T2* metrics over the course of a menstrual cycle, while anovulatory control subjects displayed no significant changes over time. T2*S metrics were significantly shorter in the pre-ovulatory phase, compared to the post-ovulatory phase. These findings suggest that an increase in collagen bound water (T2*S) is present within the ACL during the pre-ovulatory phase. Previous studies have associated shifts in tissue water content with altered mechanical properties and decreased T2* has been demonstrated in preclinical models of cyclic loading prior to gross disruption of collagen fibrils^{10, 11}. Subsequent changes in ligament stiffness over the course of the menstrual cycle may alter proprioceptive sense and contribute to increases in ACL-injury risk and laxity within the pre-ovulatory phase of the menstrual cycle.

CONCLUSION:

This is the first study to evaluate changes in ACL T2* metrics throughout the menstrual cycle. These data suggest that significant differences in ACL water content are present between pre- and post-ovulatory phases of the menstrual cycle, and may be indicative of a new imaging biomarker for ACL-injury risk.

References

1. Oiestad 2010

2. Potter 2012

3. Lohmander 2007

4. Beynnon 2014

5. Beynnon 2006

6. Shultz 2005

7. Beynnon 2015

8. Pauli 2012

9. Diaz 2012

10. Jerban 2017

11. Koff 2014

12. Juras 2013