Study design and participants
This prospective randomized controlled trial was carried out on 135 patients diagnosed as having symptomatic lumbar disc prolapse. Patients were randomly assigned into one of three groups; the first group received transforaminal injection of Magnesium sulphate with steroids (45 participants) (Magnesium sulphate group), the second group received transforaminal injection of Ozone with steroids (45 participants) (Ozone group), and the third group received transforaminal injection of steroids alone (45 participants) (Control group). Randomization was performed using the opaque closed envelope technique where the clinician picked up a sealed envelope containing a sheet of paper with the name of the group to which the patient was randomly selected. Whichever group was written on the sheet, the patient was scheduled to it. The patients were recruited from the neurology and pain clinics of Beni-Suef University Hospital, in the period from November 2020 to November 2021. The study was registered in ClinicalTrials.gov on 24/9/2020 and this is the identification number NCT04562493.
The study included patients with clinical evidence of disc bulge in the form of lumbar disc related radicular pain (radiating pain in the lower limb that follows a dermatomal pattern) of >3 months duration, not responding to conservative treatment (medical treatment and physiotherapy) and interfering with daily activities. The selected patients must have radiological evidence of posterolateral lumbar disc bulge by MRI lumbosacral.
The following patients were excluded from our study: patients with a history of spinal trauma, spinal surgery or spinal deformities, patients with radiological evidence of any inflammatory or neoplastic lesion affecting the vertebral column, spinal cord or the surrounding soft tissue, patients with clinical or radiological evidence of hip osteoarthritis, lumbar zygophysial joint arthritis [localized pain over the lumbar vertebrae that worsens with standing or bending backward and is typically relieved by bending forward], or sacroiliitis [low back or buttock pain with typical increase in severity at night and associated stiffness upon awakening, with some amelioration after exercise, and one of the following provocative tests should be positive: pelvis rock test, FABERE (Flexion, ABduction, External Rotation, Extension), and Gaenslen maneuvers]. We also excluded patients with severe lumbar disc prolapse causing lower limb weakness or sphincteric troubles, and patients with contraindications to interventions (sepsis, coagulopathy, or allergy from the used drugs). Pregnant patients were also excluded from our study. We demonstrated in a flow diagram for the recruited patients that 195 patients were assessed for eligibility. Sixty patients were excluded (28 patients didn’t meet inclusion criteria, 12 patients declined to participate, and 20 patients were excluded due to other reasons). Forty-five patients in each group received allocated intervention. Nine patients in Mg sulphate group, seven patients in Ozone group, and 10 patients in control group lost to follow-up (Fig. 1).
History was taken from the selected patients regarding the demographics and the duration of lumbar disc related radicular pain. The imaging findings regarding the number of prolapsed discs and the degree of the most prolapsed disc were also obtained.
Assessment of pain intensity and functional disability
Assessment of the pain intensity and functional disability was done before and 2 week, 1, 3, and 6 months after the interventional procedure by a neurologist who was blinded to the type of intervention. Assessment was done using Numeric Rating Scale (NRS), Oswestry Disability Index (ODI), and Functional rating index (FRI).
NRS is a single 11-point numeric scale for assessment of intensity of pain. It ranged from 0 to 10, where 0 indicates no pain and 10 indicates the worst pain .
ODI is a questionnaire used to quantify the disability from low back pain. It includes the following items: pain intensity, the level of disability of personal care, lifting, walking, sitting, standing, sleeping, traveling, sexual and social life. Each question is scored on a scale of 0–5 with 0 representing no limitation, and 5 representing maximal limitation. The scores for all questions answered are summed, then multiplied by two to obtain the index (range 0 to 100) .
FRI is a self-reported scale containing 10 items. Eight items focus on daily activities affected by back pain and 2 items focus on frequency and intensity of pain. Each item has a 5-point scale (0 = no pain, and 4 = worst possible pain). The FRI scores ranged between 0% (no disability) and 100% (severe disability) .
Assessment of patients’ satisfaction
Assessment of patient’s satisfaction about the intervention was done 6 months after the interventional procedure by a psychiatrist who was blinded to the patient’s condition and the type of intervention. It was done by using the Short Assessment of Patient Satisfaction (SAPS) scale. SAPS consists of the following seven items: satisfaction with treatment, explanation of the treatment outcome, medical care, respect by the physician, participation in medical decision making, time with the physician, and satisfaction with clinic/ hospital care. Responses scales are 5-point scales. SAPS scores were interpreted as follows: 0 -10 = very dissatisfied, 11 - 18 = dissatisfied, 19 - 26 = satisfied, and 27 - 28 = very satisfied .
Interventional pain procedure
We asked the patients to stop any medical treatment for the lumbar disc related radicular pain one week before the intervention. The selected patients were brought to the preparation room where reassurance was done. Intravenous midazolam 0.2 mg/kg was given to them, then they were placed in the prone position on fluoroscopy table and draped in a sterile manner. They were connected to a monitor (SPO2, NIBLP, and ECG) and given supplemental oxygen through a nasal cannula (3 L/min) to maintain an oxygen saturation. A 22-gauge, 3.5-inch spinal needle was used in the injection procedure. With each insertion of the spinal needle, 1 ml with 20 mg of local anesthetic lidocaine 2% was injected intradermally. Upon final needle-tip position, antero-posterior (AP) and lateral views of the fluoroscopic imaging were obtained to confirm needle positioning then the contrast was injected. AP and lateral views of the contrast spread were taken during the procedure to confirm appropriate spread into posterior epidural space.
The patients were randomly assigned into one of three groups:
Transforaminal injection of Mg sulphate:
In this group, the patients received transforaminal epidural injection of steroids and Mg sulphate with local anesthetic (7 mg Betamethasone, and 100 mg of magnesium sulphate diluted to 2 milliliters total volume with preservative-free normal saline, preceded by a test dose of 1 milliliter 2% lidocaine).
Transforaminal injection of Ozone:
In this group, the patients received transforaminal epidural injection of steroids (7 mg Betamethasone, preceded by a test dose of 1 milliliter 2% lidocaine) with the addition of an O2-O3 mixture, with an ozone concentration of 25 microgram /mL. We injected 3 mL of O2-O3 at transforaminal level.
Transforaminal injection of steroids alone:
In this group, the patients received transforaminal epidural injection of steroids with local anesthetic (7 mg Betamethasone preceded by a test dose of 1 milliliter 2% lidocaine).
After the intervention, the patients were not prescribed any medications (apart from NSAIDs for the first 3 days after injection) or specific physiotherapy regimens because they were already not responsive to them before the intervention.
Oxidative stress biomarkers (superoxide dismutase (SOD) and Glutathione (GSH)) were measured for all included patients in the three groups before and 2 weeks after the interventional procedure. For this purpose, the two-site sandwich ELISA method were employed. GSH and SOD levels were determined by reading the color changes compared to the standard curve in photometry. Samples were withdrawn from patients and the collected whole blood was refrigerated at 4 c for the night, then centrifuged it for 10 min at 1000-3000 rpm. The supernatant was taken and frozen at -80 c (for 1-3 months ) for storage.
Both markers SOD and GSH were measured by using double sandwich ELISA technique. The pre coated antibodies were human SOD and GSH monoclonal antibodies and the detecting antibody was biotin labeled polyclonal antibody. We added samples and biotin labeled antibody in ELISA wells then washed with TBS or PBS. We added Avidin peroxidase conjugate to the wells. We used TMB substrate for coloring after reactant thoroughly washed out by PBS or TBS. In presence of peroxidase activity TMB substrate changed into blue color. Finally, we added stop solution which changed blue color into yellow color. We measured the optical density (OD) by using ELIZA reader. The depth of the color and the testing factors in samples are positively correlated with their concentration. The standard curve was used to detect the amount of the biomarkers by plotting the average O. D for each standard against the concentration and drew a best fit curve using graph paper or statistical software analysis.
Because our study was the first study to compare the therapeutic effect of transforaminal injection of Magnesium sulphate versus Ozone on the oxidative stress biomarkers in patients with lumbar disc prolapse, we calculated the sample size based on the results of a pilot study we performed before starting our study. The sample size calculation was done using G*Power version 22.214.171.124 Software. The probability of type I error (α) was 5%, effect size = 0.529, df =88, critical t= 1.66, noncentrality parameter λ= 2.51, A total sample size of 45 patients in each group was required to achieve a statistical power (1–β) 80%.
IBM SPSS (Statistical Package of Social Science) Version 25 was used to analyze the data. Categorical variables such as sex and the degree of the most prolapsed disc, were expressed as numbers and percentages. Quantitative variables such as age, duration of pain, the number of prolapsed discs, NRS, ODS, FRI, SAPS, SOD and GSH were expressed as mean and standard deviation. Chi-squared test was used for comparison between Mg sulphate, Ozone, and control groups in categorical variables, whereas One-way ANOVA was used for comparison between Mg sulphate, Ozone, and control groups in quantitative variables. Paired sample t- test was used for comparison between quantitative variables before and after the interventional pain procedure. Mixed ANOVA test was used for comparing quantitative variables before and after the interventional pain procedure in Mg sulphate, Ozone, and control groups. P-value ≤0.05 was considered statistically significant. All tests were two-tailed.