**4. Discussion**

Patients with HD produce a mutant form of the huntingtin protein. This protein is widely expressed in the brain, but also in peripheral tissues [15]. Consequently, peripheral manifestations of HD have been hypothesized to be caused by cellular dysfunction caused by expression of the mutant huntingtin protein in those tissues, independent of the known neurodegenerative process of HD. However, expression of the huntingtin protein is undetectable in the heart [15] and mouse models of HD have revealed changes to the cardiovascular system in the absence of mutant huntingtin aggregates in cardiac tissues, even at end stages of the disease [9]. Therefore, measured changes of physiologic markers of ANS function are more likely to be a secondary marker of neurodegeneration affecting central pathways in areas such as the CAN. Here, we have demonstrated for the first time that patients with JOHD have significant changes in physiologic measures of ANS function compared to healthy controls. Importantly, we hypothesize that these changes are a direct consequence of pathologic changes that occur in the central nervous system of patients with JOHD rather than peripheral manifestations of the disease.

Similar to previous reports of enhanced sympathetic tone in patients with AOHD, patients with JOHD had elevations in their rHR compared to healthy controls. However, the patients with JOHD had decreases in their SBP and DBP relative to the control group, which was unexpected. It is

unclear what is driving the observed decrease in blood pressure in JOHD, but we hypothesize that earlier neurodegenerative changes may affect regions within the CAN that impact rHR. As patients progress through their disease, we believe that brain regions that are more closely related to blood pressure control are impacted. For example, the medulla oblongata receives afferent signals from the baroreceptors and affects blood pressure control [16]. The medulla has not been widely described as a primary area of neurodegeneration in HD using neuroimaging techniques. However, a post-mortem analysis demonstrated degeneration in areas of the brainstem in patients with HD [17]. This seems to fit with our hypothesis because neuroimaging studies are typically conducted in patients who have not reached the end stages of the disease, but post-mortem analyses obviously would mostly represent patients with end-stage disease, similar to patients with JOHD. To further investigate this theory, we conducted an unplanned analysis to investigate the relationship between disease duration and physiologic measures of cardiac function in the JOHD group. We found no significant relationship between disease duration and rHR (*p* = 0.936; Figure 2a). This supports the notion that rHR becomes significantly elevated early in the disease course of JOHD but does not seem to continue to worsen as the disease progresses. However, there were significant, negative correlations between disease duration and SBP (*t* = −2.20, *p* = 0.037; Figure 2b) and DBP (*t* = −2.13, *p* = 0.044; Figure 2c). These results further confirm our hypothesis that rHR is impacted earlier in the disease process of JOHD and changes in BP may be indicative of later-stage neurodegeneration.

**Figure 2.** (**A**) Disease duration among patients with JOHD did not significantly predict rHR. (**B**) Systolic BP and (**C**) diastolic BP become significantly more decreased as disease duration increased in the JOHD group. Bpm: beats per minute; DBP: diastolic blood pressure; rHR: resting heart rate; SBP: systolic blood pressure.

This analysis is the first report (to the best of our knowledge) of physiologic markers of cardiac function being disrupted in JOHD. These results come from one of the largest datasets of patients with JOHD in the world. Despite this, there are important limitations to our work. As noted above, these results only allow us to report associations but are not meant to demonstrate a causative relationship between rHR and SBP in JOHD. Similarly, the Kids-JOHD and Kids-HD studies were not focused on cardiovascular measures. While all participants had their vital signs collected in a similar setting with similar equipment by trained medical professionals, confounding factors related to the collection of rHR and SBP could have been present and may have affected these results. Additionally, we recognize that rHR and SBP are surrogate measures of ANS function. Therefore, future studies focused on investigating ANS function in JOHD should collect more precise measures, such as heart rate variability or baroreflex sensitivity. Another limitation is the potential influence of medication use on these results. We have attempted to control for this confounder by including the use of medication that increase or decrease blood pressure as a covariate in all models. However, this may not have adequately controlled for the impact of medications. To further ensure that medication use was not significantly influencing these results, we performed an unplanned sensitivity analysis where we repeated the primary analyses in participants who were not taking a medication that is known to impact blood pressure. After doing this, the participants in the JOHD group still had a significantly elevated rHR and a significantly lower SBP compared to the GNE group. Interestingly, the JOHD group also had a significantly lower DBP compared to the GNe group in this analysis. While medication use is still an important confounder, this sensitivity analysis seems to indicate that the use of these medications is not significantly impacting the reported results. It is important to note that we hypothesize that the measured changes in rHR, SBP, and DBP in patients with JOHD is mediated by CNS alterations given that JOHD is a neurodegenerative disease. However, further research is needed to support this hypothesis. It is possible that the observed changes are mediated by the peripheral nervous system, the cardiovascular system (including physical fitness), and metabolic rate. Further studies are required to determine the root cause of autonomic dysfunction in JOHD. Lastly, it is important to note that the lower blood pressures in the JOHD group were at rest. Previous reports in patients with Parkinson's Disease and Multiple System Atrophy performed orthostatic tests to determine whether patients had a precipitous drop in their BP when going from lying down to standing up, which is more indicative of autonomic function. The present study did not perform any specific measures that were meant to perturb the autonomic nervous system. Therefore, the patterns identified at rest are only theorized to be associated with central changes that could a ffect the cardiovascular system.
