Role of BoNT-A in patient with post COVID-19 spasticity, assessed by H-reflex

Background. COVID-19 is a pandemic disease due to viral infection. It spread throughout the world, and the association of COVID-19 with neuroendocrine system is a well-known fact especially while the connection between the endocrine and immune system neuroinflammation play a crucial role in pathophysiology. Therapeutic indication of Botulinum-neu - rotoxin-A (BoNT-A) is used in the treatment of focal post-stroke spasticity (PSS) and in re-habilitation which is regarded as a safe and effective type of treatment. Objective . To assess spasticity after COVID-19 infection. Methods . Thirty patients diagnosed as post-COVID-19 stroke spasticity were enrolled, their age ranged from (30-60 years). They had focal spasticity of lower extremities. BoNT-A was given in day 0 and 4 weeks later, in the gastrocnemius and soleus muscle. H-reflex was recorded from soleus muscles at 1st presentation and 4 weeks after injection of BoNT-A. Muscle power was assessed by Medical Research Council scale (MRC) and MAS. Results. The amplitude and latency of H-reflex record from soleus muscles and the H/M ratio showed a statistically signif - icant difference between pre-and post-therapy with BoNT-A injection in cases with post COVID-19 spasticity. There was a significant increase in MRC scale but, MAS scale showed a significant reduction after injection of BoNT-A. Conclusion . Post COVID spasticity is a well-known complication after COVID-19 infection, there was clinical improvement in PSS post BoNT-A injections, which was assessed clinically by MAS and MRC scale, neurophysiological the H-reflex was correlated negatively regarding latency, while there was no correlation regarding H-reflex amplitude or Hmax/Mmax ratio and MAS.


INTRODuCTION
COVID-19 pandemic is an infectious illness that spread throughout globally, which caused massive health problems [1].The association with neuroendocrine system is a well-known fact especially the connection with endocrine and immune system neuroinflammation play a crucial role in the pathophysiology, different neurological complications occur after COVID infection with 28-38% spasticity post-stroke prevalence [2].
Stroke is one of the post COVID-19 complications, and elderly stroke is one of the reasons for disability and mortality.Spasticity as a squally in those patients can be managed regardless of the cause [3].
Patients with (PSS) frequently suffer from pain, stiffness of joint, soft tissue that leads to contracture with abnormal posture of extremities [4].Limitation in joint movement was estimated to occur after stroke within 3-6 weeks [3].
Early diagnosis and treatment of PSS would reduce the complications, improve functions of joints muscles and extremities with better life style for such patients [5,6].
The BoNT-A injections are used clinically for treatment due to its safety and effectiveness for patients with spasticity [7,8].They reduce spasticity which improves muscle function, and range of movement in the lower extremities [9,10].The actions of BoNT-A injection are inhibition of exocyto-sis of acetylcholine (Ach) from presynaptic nerves terminals on the neuro-muscular junctions [11].It acts by inhibition of selective reversible muscle contraction with no undetectable general weakness and sedation lasting for three to four months [12].
The H-reflex is used to study neuro-motor control processes and clarifying neuromotor deficits [13,14].Previous work revealed that patients suffering from spasticity had a larger amplitude of H-reflex and Hmax/Mmax ratio than normal subjects or the un-diseased side of patients at resting position [15], which means spasticity at rest is due to motor neurons hyper excitability of spinal circuits.The measurements H-reflex gives a clue of how effective the therapeutic interventions are (botulinum toxin) on reflex pathways [16].
This study aims to explore H-reflex changes in survivors with post covid spasticity and the effect of BoNT-A on PSS, and to assess the correlations if there are any regarding the changes in H-reflex with (MAS) Scale and (MRC) scale after treatment with BoNT-A injections.

METHODS
A cohort prospective research, which was carried out at the Clinical Neurophysiology Unit, Emamein Kadhimein Medical City from January to August 2023.Each patient of the study had signed a consent for participation.
The study involves 30 patients of both sexes, age ranged from 30 -60 years.The 14 males and 16 females were referred by a specialist Neurologist.
Patients diagnosed with COVID-19 complicated by stroke had post stroke spasticity.They were referred from the specialist after a thorough clinical and neurological examination with assessment of muscle tone of ankle joints by the use of modified Ashworth Scale (MAS) [17,18], muscle power of the lower extremities was tested according to the Medical Research Council (MRC) scale [20], the examination was done at day zero then 3-4 weeks later.The diagnosis was confirmed by CT scan.
Exclusion criteria of patients were: Any neurological or neurodegenerative disease, orthopedic or musculoskeletal disorder, any disease or medications (like muscle relaxant) that affect the tests of nerve conduction, lastly patients in whom BoNT-A is contraindicated.
Patients were received in the early morning, H-reflex testing was done at day zero and then it was repeated 3-4 weeks later, the test was done by EMG/EP machine (Medtronic Keypoint, Denmark).The temperature of the room was monitored around 25º C to 28° C through the test procedures, the temperature of skin was measured by skin thermometer to keep it between 32 -34° C.
H-reflex was recorded from the soleus muscles.The onset of M-latency and H-reflex were calculated from the starting point of stimulus artifacts to the first deflection from the baselines.The amplitudes of H-reflex, M-wave were measured as peak-to-peak value.Post recording of the H-wave then stimulus strength was adjusted to find Hmax and Mmax slowly, and the Hmax/Mmax ratio were measured (the maximum amplitudes ratio of the two action potentials) [19].
All patients involved in the study received an injection of BoNT-A toxin at: day zero then 3-4 weeks later.The vial of BoNT contains 100U which was diluted according to the method of El-Tamawy et al. [20].All patients were injected with (100 U/session).The target muscles injected were gastrocnemius, soleus according to palpation/anatomic landmarks of International Movement Disorder Society guidelines [21].
Results are expressed as mean standard deviation using Microsoft Excel 2016 and IBM SPSS version 26.Continuous pre-and post-treatment results were tested by paired student (t) test, while median and range was used for Non-continuous variables using the Wilcoxon test for comparison.A p-value < 0.05 was considered significant.
Table 1 shows: The MRC scale scoring was; 4 patients grade I, 14 cases grade II, while 12 cases were grade III and zero patient grade IV (Before treatment), while After treatment it was:0,1,9 and 20 respectively with a significant (P= 0.001).
Regarding MAS scale Score; 16 of patients had score 4, 8 of them had 3 score, while 5 patients scored 2, lastly one of them had a score of 5.
There was significant increase in MRC scale score (p<0.001)post-treatment compared to pre-treatment value, in contrary there was significant reduction in MAS scale after treatment with Bot-A with a (p <0.001),Table 1.The data of H-reflex in right and left sides of lower extremities: There was prolongation in latency while there was a decrease in amplitude and the H/M ratios after injections compared to patients before therapy (Table 2).
Following BoNT-A injection (Table 3) shown the duration of H-reflex was related negatively with MAS (r = -0.286;p = 0.044) as shown below.The association between the H-reflex data and sex in Table 4; there was no significant association in different sex.

DISCuSSION
In this study, treatment with the injections of BoNT was found to have the ability to reduce spasticity of paretic side.There was a decrease in MAS scores in all cases with variable degrees that indicated that BoNT was effective in reducing spasticity and which dose was adequate for these subjects.Similar results were found by Lopez et al in 2019 [22].The effects of Botulinum toxins are related to the inhibition of Ach releasing at the neuro-muscular junctions [23], leading to a local neuromuscular block at site of injection, this re- sulted in paresis of the targeted spastic muscles, so it will reduce spasticity in the injected muscle with BoNT.
There was a significant elevation in cumulative MRC score at 2nd visit post injection with BoNT-A; which means an increase in muscle strength [24].
Spasticity is a UMN positive sign; because there is increase in muscle tone and stretches reflex, while weakness and decrease in muscles strength are considered the "negative" UMN sign [25].These cycles of positive and negative motor neuron lesions are one of clinical complains in the recovery period post-stroke [26,27].Consequently, contracture resulted due to the weakness and spasticity leading to immobilizations of the joint at a shortened muscles length, these contractures will exacerbate spasticity in muscles.Such a vicious cycle continues and the patient's condition becomes worse unless treated effectively [28,29].
The decrease in spasticity of muscles is due to the reduction of reciprocal inhibitions from the antagonistic muscles post injections.Different studies have showed that injection can paralyze afferent fibers [30,31], as well as the blockage of ACh release from pre-synaptic terminal at neuro-muscular junction.
There was a good degree of increment in muscle strength simultaneous with decreased spasticity as assessed by "MRC scale and MAC, respectively after BoNT-A injection were of value for those patients with PSS".
The values of H max /M max ratios were significantly greater in the affected limb of stroke cases before the injection of BoNT-A due to higher excitability of moto-neuron pool in such patients (Phadk CP 39).After BoNT-A injections, the amplitudes and Hmax/Mmax ratio were decreased in the limb with spasticity of stroke cases.This suggests a decline in the motoneuron pools of excitability.
Hyper-excitability of stretch reflexes are the pathophysiological changes that occur in spastic patients after stroke in patients with PSS, with alteration in the descending conductions pathway above the spine levels which may affect the excitability of alpha motor neurons [32].The increased excitability of alpha-motor neurons may represent the severity of spasticity.The excitability of alpha motor neurons is detected in this study by using H-reflex; to reflect and to assess the peripheral sensory afferent or supra-spinal-descending conductions pathway in various aspects of human movements.Our patients with PSS had fewer latency of H-reflex.Joodaki et al., in 2001Cecen et al. in 2018 had similar observations [32,33].After BoNT-A injections, our data showed prolongation in the latency of H-reflexes.
In this study, there were negative correlations between latency of H-reflex and MAS post injections of BoNT-A.These findings of the relation between H-reflex and MAS would make the H-reflex and MAS as clinical and neuro-physiological tools for assessment of muscle spasticity.The finding of no relation between MAS and H max /M ma might be related to the fact that Hmax/Mmax ratios reach the maximum between 8 weeks -24 weeks and regardless weather H-reflex can be elicited as short as post the spinal cord injury [34].So, it`s recommended to examine cases at least 6 months after the disorder process.
However, studies on post-stroke spasticity patients regarding neurophysiological examination had mainly focused chronic cases [35][36][37].Since the average time for spasticity to occur is 1 month after the onset of stroke and abnormal neuroplasticity develops [38], early identification of spasticity is necessary for early and adequate treatment and hence there are better outcomes.In parallel with the neuro-physiological results, a recent meta-analysis suggested that post-stroke spasticity usually appeared or disappeared within one to three months post-stroke and remained stable beyond three months [39].
In this study, no correlation was found between the H max /M ma ratio and the MAS scores.This could be attributed to the mechanisms underlying spasticity where there are changes in spinal interneurons and motor neuron excitability or could be due to mistakes inherent to the MAS scale, which rely on the subjective judgment of the investigator, measuring the resistance to passive muscles stretching.This resistance often reflects the combination of spasticity, thixotropy and fixed muscle contractures.A recent study on the evolutional changes in neurophysiology of the spinal segmental circuitry has showed that the amplitudes of flexor reflexes may fall as spasticity becomes established [40][41][42].
Here, there was exclusion of cases with clinically detectable fixed contracture, but it's not possible to exclude the changes which may occur in muscle fiber structures or the visico-elastic features.
This study has some limitations that need to be addressed.It examines the relationship between MAS and H-reflex parameters in a moderate sample size (n = 30).A great sample size is recommended to evaluate the changes in neuro-physiological tests.The duration of the study could be prolonged to evaluate the effects of BoNT-A injection on MAS and H reflex in chronic PSS patients.

CONCLuSION
This study provides evidence that patients with post COVID spasticity have clinical improvement after BoNT injections assessed by MAS and MRC scales, in addition to significant differences regarding the latency and amplitudes of H-reflex in those patients preand post-BoNT-A injection.

Acknowledgments:
We would like to thank everyone who participated in this work starting from patients, all the neurophysiology department in Al-Emamein Kadhimein Medical City including physicians, nurses and specialists.Contribution of authors: Dr. Safa has done the clinical examination of patients and performed the H-reflex test for them and the injection of BoNT-A, Dr. Israa has put the plan of the study, collected the data analyzed it, made the tables and wrote the article.Dr. Safa and Dr. Islam had collected the patients, Dr. Safa did the neurological tests, Dr. Israa had put the plan and wrote the paper.

Conflict of interest: none declared
Financial support: none declared

TABLE 1 .
Muscle strength and tone scoring according to MRC scale and MAS of the studied group (n=30)

TABLE 2 .
H-reflex data before and after treatment with-BoNT-A

TABLE 3 .
Findings of H-reflex correlation with; MRC, and MAS after BoNT-A treatment

TABLE 4 .
Association between different sex (male and female) with the Data of H-waves and M-waves