**3. Results**

Participant characteristics are shown in Table 1 and Supplemental Table S1. There were 154 men and 214 women, with an average age of 63.8 ± 10.5 years. There were 156 (76 male; 80 female) and 212 (78 male; 134 female) participants in the N and L groups, respectively.

#### *3.1. Adult Participants*

The average age of the adult group was 54.3 ± 7.3 years. In total, 71 (28 male; 43 female) and 92 (24 male; 68 female) participants were included in the N and L groups, respectively (Table 2, Supplemental Table S2). The body fat percentage (BFP) was significantly higher and the grip strength and gait speed were significantly lower in the L than in the N group (BFP; N: 28.2 ± 5.1, L: 31.0 ± 6.8, *p* = 0.005, grip strength; N: 29.6 ± 9.8, L: 25.1 ± 8.7, *p* = 0.002, gait speed; N: 2.4 ± 0.3, L: 2.3 ± 0.5, *p* = 0.04).

Analyses of the laboratory data and nutrient intakes indicated that LDL-C was significantly lower and Vitamin B1 was significantly higher in the L than in the N group (LDL-C; N: 132.1 ± 32.4, L: 121.8 ± 28.5, *p* = 0.033, Vitamin B1; N: 0.66 ± 0.07, L: 0.69 ± 0.09, *p* = 0.029). There were no significant between-group differences in the other laboratory data, nutrient intake, SMI, or history of metabolic diseases (Table 2, Supplemental Table S2).

As significant differences were observed among several factors, they were examined as covariates for risk factors of the L group in the logistic regression analysis. BFP and LDL-C were risk factors (BFP; exp (B) 1.068, 95% CI: 1.003–1.127, *p* = 0.038, LDL-C; exp (B) 0.988, 95% CI: 0.977–1, *p* = 0.042) (Table 3).

#### *3.2. Older Adult Participants*

The average age of older adult participants was 71.3 ± 5.3 years. The N and L groups contained 85 (48 male: 37 female) and 120 (54 male: 66 female) participants, respectively (Table 4, Supplemental Table S3). The average age of participants was significantly higher in the L than in the N group (N: 70.2 ± 4.7; L: 72.2 ± 5.6, *p* = 0.008). The BFP was significantly higher and the grip strength was significantly lower in the L than in the N group (BFP; N: 26.7 ± 6.3, L: 29.7 ± 6.9, *p* = 0.002, grip strength; N: 29.0 ± 8.0, L: 25.1 ± 8.0, *p* = 0.001).

Analyses of laboratory data indicated that Hb, albumin, and calcium levels were significantly lower in the L than in the N group (Hb: 13.8 ± 1.0, L: 13.4 ± 1.1, *p* = 0.036, albumin; N: 4.4 ± 0.2, L: 4.3 ± 0.2, *p* = 0.030, calcium; N: 9.2 ± 0.3, L: 9.1 ± 0.3, *p* = 0.025). Regarding nutrient intake, sodium, MUFA, and n-6 PUFA were significantly higher in the L than in the N group (sodium: N: 1961.1 ± 586.7, L: 2183.1 ± 789.0, *p* = 0.029; MUFA: N: 15.8 ± 3.8, L: 17.2 ± 5.4, *p* = 0.047, n-6 PUFA; N: 10,898.3 ± 2808.4, L: 12,149.4 ± 4807.4, *p* = 0.0233). The prevalence of hypertension was higher in the L group than in the N group (N: 42.3%, L: 60.8%, *p* = 0.007). There were no significant between-group differences in gait speed, SMI, other laboratory data, nutrient intake, or history of metabolic diseases between the N and L groups (Table 4).

Factors for which significant between-group differences were observed were examined as covariates for the risk factors of the L group in the logistic regression analysis. BFP, sodium, and hypertension were risk factors (exp (B) 1.073, 95% CI: 1.017–1.132, *p* = 0.011, sodium; exp (B) 1.001, 95% CI: 1–1.001, *p* = 0.032, hypertension; exp (B) 2.288, 95% CI: 1.186–4.412, *p* = 0.014) (Table 5).


**Table 1.** The comparison of each parameter between nonolder adult and older adult participants.

Values are expressed as means ± standard deviations; BMI: body mass index, BFP: body fat percentage, SMI: skeletal muscle mass index, y/n: yes/no; SMI: skeletal muscle mass index, N/L: normal group/locomotive syndrome group, y/n: yes/no; HDL-C: high-density lipoprotein cholesterol, LDL-C: low-density lipoprotein cholesterol; SFA: saturated fatty acid, MUFA: monounsaturated fatty acid, PUFA: polyunsaturated fatty acid, HUFA: highly unsaturated fatty acid. All analyses are comparisons between adults and older adults. The comparison of male/female was conducted by a chi-square test, and the comparisons of the others were conducted by a Student *t*-test. \*: *p* < 0.05.

**Table 2.** The comparison of each parameter between the N and L groups in adult participants.



**Table 2.** *Cont.*

Values are expressed as means ± standard deviations. BMI: body mass index, BFP: body fat percentage, SMI: skeletal muscle mass index, y/n: yes/no, HDL-C: high-density lipoprotein cholesterol, LDL-C: low-density lipoprotein cholesterol, SFA: saturated fatty acid, MUFA: monounsaturated fatty acid, PUFA: polyunsaturated fatty acid, HUFA: highly unsaturated fatty acid. All analyses are comparisons between the N and L group. The comparisons of male/female, hypertension, diabetes, and hyperlipidemia were conducted by a chi-square test, and the comparison of the others were conducted by a Student *t*-test. \*: *p* < 0.05. There were significant differences in BFP, grip strength, gait speed, LDL-C, and Vitamin B1 between the N and L groups.

**Table 3.** Logistic regression analysis for risk factors of the locomotive syndrome (L group) in adult participants.


BFP: body fat percentage, LDL-C: low-density lipoprotein cholesterol. \*: *p* < 0.05. Covariates: BFP, grip strength, gait speed, LDL-C, Vitamin B1. There were significant differences in BFP and LDL-C.

**Table 4.** The comparison of each parameter between the N and L groups in older adult participants.



**Table 4.** *Cont.*

Values are expressed as means ± standard deviations. BMI: body mass index, BFP: body fat percentage, SMI: skeletal muscle mass index, y/n: yes/no, HDL-C: high-density lipoprotein cholesterol, LDL-C: low-density lipoprotein cholesterol, SFA: saturated fatty acid, MUFA: monounsaturated fatty acid, PUFA: polyunsaturated fatty acid, HUFA: highly unsaturated fatty acid. All analyses are comparisons between N and L group. The comparisons of male/female, hypertension, diabetes, and hyperlipidemia were conducted by a chi-square test, and the others were conducted by a Student *t*-test. \*: *p* < 0.05. There were significant differences in age, BFP, grip strength, hemoglobin, albumin, calcium, sodium, MUFA, n-6 PUFA, and hypertension between the N and L groups.

**Table 5.** Logistic regression analysis for risk factors of the locomotive syndrome (L group) in older adult participants.


BFP: body fat percentage, MUFA: monounsaturated fatty acid, PUFA: polyunsaturated fatty acid, \*: *p* < 0.05. Covariates: age, BFP, grip strength, hemoglobin, albumin, calcium, sodium, MUFA, n-6 PUFA, and hypertension. There were significant differences in BFP, sodium, and hypertension.

#### **4. Discussion**

There have been several reports on the relationship between motor function and muscle and nutrition [10,11]. However, to our knowledge, no study has directly examined the relationship between nutritional intake status and LS. In this study, salt, MUFA, and n-6 PUFA intakes were significantly higher in group L among older adults. Multivariate analysis indicated that higher salt intake was particularly associated with LS.

In an aging society, the rate of LS is increasing, and the number of people who have difficulty leading independent daily lives is increasing [4]. Therefore, preventing LS plays a vital role in preventing the need for nursing care. In this study, a significant decrease in grip strength was observed in the L group in both adults and older adults. This suggests that a decline in muscle strength accompanies the functional decline in LS and that the influence of muscle strength on LS is significant and closely related to sarcopenia. Exercise and nutrition are important factors in preventing sarcopenia [32] and are thought to be equally important in influencing LS. Exercise therapy has been advocated for LS prevention [9]. However, there have been no detailed studies on the effects of nutrition on LS. Thus, there is no specific diet therapy for LS prevention.

BFP and LDL-C levels were lower in group L among adults, according to the multivariate analysis. Higher body fat may reflect lower physical activity, such as lack of exercise, in group L. High LDL cholesterol is associated with a higher percentage of fat than muscle [33]. However, in this study, LDL-C levels of both groups were within the normal range, so the difference was not clinically significant.

Vitamin B1 intake was lower in group L among adults, according to the univariate analysis. Vitamin B1 is active in glucose metabolism, and its deficiency can lead to poor conversion of sugar to energy and increased fatigue [34]. In this study, the intake of Vitamin B1 was higher in group L, but both groups had deficient Vitamin B1 intake compared to the daily recommended amount [35], and the difference in intake was small and not significant for LS.

Among older adults, group L had higher salt intake and hypertension, according to the multivariate analysis. Excessive salt intake is one of the causes of various diseases, such as hypertension [36,37], where sodium in the skeletal muscle accumulates more in older than in younger people, and patients with refractory hypertension have increased tissue sodium (Na+) content compared to normotensive controls [38]. Moreover, the sodium-potassiumchloride symporter 1 (NKCC1) is highly expressed in mammalian skeletal muscles. The physiological function of NKCC1 in myogenesis remains unclear. However, NKCC1 protein levels increase skeletal myoblast differentiation, and NKCC1 inhibitors markedly suppress skeletal myoblast differentiation [39]. It has also been reported that excess sodium leads to downregulation of NKCC expression [40]. Further, the risk of sarcopenia, which is associated with decreased muscle mass and strength, is related to the amount of salt intake because Na+ is stored in tissues, and NKCC1 is involved in muscle hypertrophy and suppression [41]. Here, salt intake was higher in the L group.

Regarding hypertension and muscle, hypertension reduces muscle blood flow [42] and correlates with low muscle mass [43]. Arterial stiffness and age-related hormonal decline are risk factors for hypertension and muscle loss [44]. Similar to these observations, this study also detected an association between hypertension and LS. Thus, higher salt intake and hypertension were associated with muscle metabolism and sarcopenia, which may be associated with the locomotive syndrome. In this study, we found that the locomotive syndrome group had weaker grip strength, which may be due to muscle weakness and sarcopenia. Therefore, it can be said that factors related to muscle mass loss are closely related to the locomotive syndrome.

In contrast, even after multivariate analysis of confounding factors involved in locomotive syndrome, including grip strength, higher salt intake was significantly associated with locomotive syndrome. Higher salt intake may also be related to motor nerves, sensory nerves that control balance, and other factors, in addition to its effect on muscle mass. There are many unclear points regarding the relationship between higher salt intake and nonmuscle factors; thus, further basic research is needed in the future.

Among older adults, there were no significant differences except for salt intake and hypertension in multivariate analysis. However, there were significant differences in MUFA and PUFA intake, hemoglobin, serum albumin, and calcium in the univariate analysis. Although there was no significant multivariate difference between n-6 and n-3 PUFA levels and locomotive syndrome, there may be a small relationship with locomotive function. In recent years, several studies have reported that n-6 and n-3 PUFAs have opposite effects on insulin resistance (IR) and body homeostasis [45]. It is postulated that n-3 PUFAs attenuate the development of IR by reducing inflammation, whereas n-6 PUFAs promote IR [46]. More recent evidence indicates that n-6 PUFAs play a key role in the inflammatory process and are associated with various metabolic diseases [47]. Therefore, it seems reasonable to control the dietary ratios of n-6/n-3 PUFAs to ameliorate obesity-related IR, which is beneficial for protection against chronic and metabolic diseases. A study of rats fed a highfat diet demonstrated that the accumulation of fatty acids in various lipid fractions increased in the gastrocnemius red muscle when there was a shift in the n-6/n-3 PUFA balance in favor of n-6 PUFAs. The increases in n-6 PUFAs are associated with fat accumulation in muscle [48]. A high percentage of n-6 PUFAs was also associated with LS in this study.

This study had several limitations. First, the participants were middle-aged and older adults who lived in a relatively rural area and were employed in agriculture or fishing. Thus, the differences in lifestyles between these participants and people in an urban environment may limit the generalizability of our results. Second, the participants attended annual health examinations, suggesting that they may be more health-conscious than other people. Third, this was a cross-sectional, single-center study. In the future, longitudinal and multicenter collaborative research is needed to verify our findings. Fourth, BIA is not the gold standard assessment for body composition (the 4-compartment model or DXA are better).

## **5. Conclusions**

In conclusion, this study examined the relationship between LS and nutrition. Salt intake and hypertension were associated with locomotive syndrome. Thus, we sugges<sup>t</sup> that limiting salt intake may effectively prevent LS.

**Supplementary Materials:** The following supporting information can be downloaded at: https: //www.mdpi.com/article/10.3390/jcm11030610/s1, Table S1: The comparison of all parameters between non-elderly and elderly participants; Table S2: The comparison of all parameters between the N group and L group in adult participants; Table S3: The comparison of all parameters between the N group and L group in older adult participants.

**Author Contributions:** Conceptualization, S.I. (Sadayuki Ito); methodology, S.I. (Sadayuki Ito); software, K.W.; validation, K.W.; formal analysis, S.I. (Sadayuki Ito); investigation, S.I. (Sadayuki Ito), H.N., K.A., M.M., T.S., S.I. (Shinya Ishizuka), Y.T., Y.H., and S.I. (Shiro Imagama); resources, S.I. (Sadayuki Ito); investigation, S.I. (Sadayuki Ito); H.N., K.A., M.M., T.S., S.I. (Shinya Ishizuka), Y.T., Y.H., and S.I. (Shiro Imagama); data curation, S.I. (Sadayuki Ito); writing—original draft preparation, S.I. (Sadayuki Ito); writing—review and editing, S.I. (Sadayuki Ito) and H.N.; supervision, S.I. (Shiro Imagama); project administration, H.N. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research did not receive any specific gran<sup>t</sup> from funding agencies in the public, commercial, or not-for-profit sectors.

**Institutional Review Board Statement:** The research protocol was approved by the Human Research Ethics Committee and the university's Institutional Review Board (No. 2014-0207). All participants gave written informed consent prior to participation. The research procedure was carried out in accordance with the principles of the Declaration of Helsinki.

**Informed Consent Statement:** All participants gave written informed consent prior to participation.

**Data Availability Statement:** The data of health check-ups used to support the findings of this study are available from the corresponding author upon request.

**Acknowledgments:** We are grateful to the staff of the Comprehensive Health Care Program held in Yakumo, Hokkaido, and Aya Henmi and Hiroko Ino of Nagoya University for their assistance throughout this study.

**Conflicts of Interest:** The authors declare no conflict of interest.
