The Fall Efficacy Scale International

Published Date: 02 Nov 2017

7.0. Introduction

Ageing represents a highly complex and inevitable process, involving biochemical, physiological and systemic alterations across numerous body systems (Xu, Knutson,Carter & Leeuwenburgh, 2008; Ryan, Dudash, Docherty, Geronills et al, 2010). Oxidative stress has been highlighted as a causative factor towards declining physical function in human skeletal muscle (Pansarasa, Castagna, Colombi et al., 2000; Fano, Mecocci, Vecchiet et al., 2001). Harman first introduced the free radical theory into the literature in 1956, postulating that with advancing age free radicals generated in normal metabolic processes are focal contributors towards cellular and tissue oxidative damage to proteins, lipids, and nucleic acids (Harman, 1956). Research interest focussing on the commonalities between inflammation and oxidative stress has increased exponentially over the past decade, this has not been exclusive to clinical setting but has extended to the fields of exercise biochemistry and immunology (Ji, 2001; Peake, Suzuki & Coombes, 2006). It has been postulated that both inflammation and oxidative stress can activate parallel intracellular signalling pathways, which have been linked to the age associated loss of muscle mass and strength (Bonetto et al. 2009).

Inflammation is a fundamental characteristic of ageing, heightened levels of inflammatory markers, including pro-inflammatory cytokines (e.g., tumor-necrosis-factor (TNF)-α, interleukin (IL)-6) are synonymous with advancing age (Franceshi, Capri, Monti, 2007). Although the etiology of increased inflammatory cytokines is multifactorial, heightened generation of free radicals and oxidative stress are believed to be central contributors (Mc Ardle & Jackson, 2011). Oxidative stress has been shown to enhance activation of the pro inflammatory nuclear factor-kB signal transduction pathway, which resultantly increases Interleukin-6 and TNF α production (6, 10). However the body contains an elaborate antioxidant defence system to protect the body from the harmful effects of heightened levels of oxidative stress, encompassing antioxidant enzymes alongside radical-scavenging antioxidants (Halliwell, 1999; Fusco et al., 2007). In situations whereby the production of oxidants exceeds the body’s scavenging and buffering ability, the intracellular redox balance can shift towards a pro-inflammatory environment (Packer et al., 2000; Ji et al., 2001; Powers et al., 2008). Consequentiality this can cause damage to proteins, DNA, and lipids, which has been shown to increase with advancing age (Lim, Cheng & Wei, 2002; Ceserai et al., 2004; Gianni, Jan, Douglas, Stuart & Tarnopolsky, 2004; Powers et al., 2008). Although research has postulated that certain dietary antioxidants, may be effective in suppressing the activation of such proinflammatory pathways through the quenching of free radical molecules (11, 12), justifying their inclusion within the elderly population.

Considering the proven links between oxidative stress, free radicals and the ageing process theoretically it would be reasonable to assume that interventions which could limit or inhibit the production of free radicals should be beneficial towards the intracellular environment (Harman, 2003; Meng et al., 2010; Mc Ardle & Jackson, 2011). Previous research conducted within the area has suggested that lower levels of antioxidants contribute towards decrements in physical function and disability (Cesari et al., 2004; Bartali, Frongillo, Bandinelli et al., 2006; Ble, Cherubini, Volpato et al,. 2006; Bartali, Semba, Frongillo et al., 2006; Bartali et al., 2008). Reduced levels of serum carotenoids have also been associated with poor muscle strength (Semba et al., 2003), in the InCHIANTI study reduced β-carotene intake was also associated with impaired lower extremity performance (Cesari et al., 2004). Whereas antioxidant supplementation offers promise in maintaining hormesis, despite increased research attention supporting evidence for its inclusion and beneficial effect is limited and largely conflicting.

The aim of this study is to examine the chronic effects of resistance exercise on inflammation, body composition and muscle strength in a healthy population, to ascertain if completion of chronic exercise alters the acute response to exercise. It is believed that individuals with greater levels of body mass will also posses higher levels of bone mineral density and muscle mass.

7.1- Methods

7.1.1-Subject characteristics

Following approval from the Office for Research and Ethics Committee in Northern Ireland (ORECNI) and the South Eastern Health and Social Care Trust, forty nine (n=13) healthy older females were recruited from the University of Ulster Alumni database, AGE NI, local walking groups and leisure centres within the area via a recruitment poster. Inclusion criteria required individuals to be community dwelling and to gain a score of 25 or above out of 30 in a Mini Mental state examination (MMSE, Folstein, 1975), as a marker of ability to provide fully informed consent. Individuals with cardiopulmonary disease or neuromuscular impairment, angina or congestive heart failure, or those on hormone therapy were excluded from participating in the study. All procedures and potential risks were fully explained prior to exercise. Each participant was provided with an information sheet and informed consent form to sign (previously outlined in methods section 3.4, See Appendix 2).

Table 7.1- Age and physical characteristics at baseline

Dependent Variable

Healthy individuals

(n=13)

Age (yrs)

70.53 ± 4.03

Stature (m)

163.50 ± 6.63

Body Mass (kg)

63.63 ± 7.20

BMI ( kg/ m2)

24.2 ± 3.11

7.1.2-Experimental Design.

Each participant ingested an antioxidant cocktail containing alpha lipoic acid, Vitamin C and Vitamin E. Each capsule contained: Alpha lipoic acid (100mg, Actual Alpha Lipoic Acid), Vitamin C (500mg, Actual Vitamin C) and Vitamin E (D-Alpha Tocopheryl Acetate, 200 IU Actual Vitamin E) daily. One tablet was consumed twice daily for 8 weeks. All testing procedures were conducted between 9.00 and 11.00 am following a standard 10 hr overnight fast to control for biological variation.

7.1.3-Anthropometric measurements

Upon arrival at the laboratory, measurements of stature and body mass were recorded for each participant as outlined in the methods section 3.11.1. Total body imaging was acquired using the GE Healthcare Lunar iDXA and analysed using enCORE software version 13.0. Total body scans were completed at baseline and post 8 week supplementation intervention to measure total body composition, bone mineral content (BMC, in g), and bone mineral density (BMD, in g/cm2). For body composition analysis, the total fat (%), total lean mass (kg), total fat mass (kg), arm fat (%), arm lean mass (g), leg fat (%), leg lean mass (g), android fat (%) and gynoid fat (%) were recorded. Daily quality control scans were acquired during the study period outlined in methods section 3.11.2. Participants were scanned using standard imaging and positioning protocols.

7.1.4. -Hand grip strength.

Hand grip strength was employed in the present study as an overall indicator of general muscle strength (Rantanen et al., 2003). Measurements of isometric grip strength were conducted using an adjustable hand held digital dynamometer (Takei Digital Grip Strength Dynamometer, Japan) in the dominant and non dominant hand. During the test the dynamometer was fixed to the arm of the chair with the elbow flexed in an angle of 90°. Test- retest reliability for hand grip has demonstrated a high correlation (r≥0.80) (Segura-Orti & Martínez-Olmos, 2011). Participants were encouraged to squeeze the handle as hard as possible. The maximal value of three trials was noted in kilograms.

7.1.5- Timed get up and go (TUG).

The TUG was completed using a standard chair, and with the use of walking aid if required. During testing procedures the individual was timed rising up from the chair, walking 3 metres to a line of tape on the floor, turning around and returning to the chair and sitting back down. Time was recorded using a stop watch, individuals completed the test pre and post supplementation intervention.

7.1.6- Fall Efficacy Scale International (FES-I).

The FES-1 was administered at baseline and following completion of 8 week supplementation

period. Individuals scored 16 questions on a 1-4 likert point scale, total score was calculated by adding each of the scores for the 16 questions (Yardley et al., 2005).

7.1.7- Collection and analysis of blood samples

Venous blood was collected during 4 separate time points, (1) pre supplementation/pre acute resistance 1, (2) post acute resistance, (3) post supplementation /pre acute resistance 2, (4) post acute resistance 2. Blood samples were drawn aseptically from an antecubital forearm vein using the vacutainer method (methods section 3.15).

7.1.8- Blood analysis.

Blood was analysed for TNFα, IL-10 and IL-6 via enzyme linked immunosorbent assay (ELISA) using the microplate reader manufacturers details. Detailed descriptions of all biochemical and ELISA assays can be found in (Methods section 3.15.2).

7.1.9-Lipid Hydroperoxide measurement.

Lipid hydroperoxides were measures spectrophometrically in serum samples using the FOX-1 assay. A detailed description of this assay can be obtained by referring to the methods section 3.15.3.

7.1.10- Measurement of lipid soluble antioxidants.

Retinol, alpha tocopherol, gamma tocopherol, beta carotene, alpha carotene and lycopene were determined by High performance liquid chromatography (HPLC) (Thurnham et al., (1998). For further description see Method section 3.18.

7.1.11- Ascorbyl radical measurement (Electron paramagnetic resonance –EPR)

Spectra were obtained using a Bruker (Model EMX Micro 6) spectrometer using a TM100 cavity and an aqueous flat cell. EDTA samples were prepared as described in greater detail in methods section 3.17.1. Experimental EPR spectra were simulated using WinEPR and Data Acquisition programs (Version 2.11, Bruker Win EPR System) and filtered identically. The average spectral peak-to-trough line height was considered as a measure of the (relative) radical concentration.

7.1.12-Statistical analysis.

Statistical analysis was performed using the SPSS social statistics package (Version 19.0, Surrey, UK). Data were analyzed using parametric statistics following mathematical confirmation of a normal distribution using Shapiro-Wilks tests. The linear relationship between the two continuous variables of interest was established using Pearson’s product moment correlation. Pre and post chronic exercise data were analyzed using an one way analysis of variance to compare means (ANOVA). Acute exercise data were also were analyzed using an one way analysis of variance to compare means (ANOVA) including 4 time points (Pre acute 1, Post acute 1, Pre acute 2, Post acute 2) with a posteriori Tukey Honestly Significant Difference (HSD) test. The alpha was established at P<0 .05 (95% confidence interval) and all values are reported as a mean ± standard deviation (SD).

7.2. Results

All participants ingested an antioxidant cocktail containing alpha lipoic acid, Vitamin C and Vitamin E, twice a day for 8 weeks. No issues were reported during the supplementation phase. For body composition analysis 13 individuals completed DXA scans pre chronic exercise and 12 individuals completed post scans. No drop outs were experienced over the time course, blood samples were all successfully completed.

Table 7.2- Physiological and physical performance measurements pre and post supplementation phase.

 

Healthy Individuals

 

Pre Supplementation

Post Supplementation

Hand grip (right) (kg)

24.72 ± 3.74

26.85 ± 3.72

Hand grip (left) (kg)

22.62 ± 3.24

24.63 ± 3.26

FES-I

18.38 ± 3.15

18.69 ± 2.59

Timed get up and go

7.52 ± 0.86

7.14. ± 0.81

Acute Resistance scores

54.30 ± 8.11

59.49 ± 8.59*

1 Repetition Maximum (kg)

59.0 ± 8.74

67.31 ± 6.06*

IL-6 (pg/ml)

4.74 ± 1.10

3.43 ± 1.18*

IL-10 (pg/ml)

4.39 ± 0.79

7.60 ± 1.88*

TNFα (pg/ml)

6.29 ± 0.41

6.55 ± 1.10

* Indicates a difference between group as a function of state (pre vs post scores) (P < 0.05).

7.2-Chronic Exercise Period

7.2.1- Hand Grip Strength- Following 8 weeks of antioxidant supplementation hand grip strength scores improved by 8.6% and 8.8% respectively as displayed in Table 7.2, however this relationship was not found to be statistically meaningful for either right or left hand grip scores (pre vs. post chronic, P.>0.05).

7.2.2-Falls Efficacy Scale- No significant relationships were found for FES-I scores between pre and post supplementation. FES-I scores experienced a slight increase post supplementation, however levels were relatively consistent during both time points (18.38 ± 3.15 vs. 18.69 ± 2.59, P>0.05).

7.2.3-Timed Get up and Go (TUG)- 8 weeks of antioxidant supplementation did not elicit a significant relationship between TUG scores. As displayed in Table 7.2 Timed get up and go post supplementation values were 5% lower than baseline values.

Table 7.3– Antioxidant and oxidative stress measures pre and post supplementation phase

 

Healthy Individuals

 

Pre Supplementation

Post Supplementation

Lipid Hydroperoxide (LOOH) µ M.L-1

0.23 ± 0.14

1.55 ± 1.11*

Gamma tocopherol (mmol· L-1)

1.5 ± 0.6

2.06 ± 0.44*

Alpha tocopherol (mmol· L-1)

19.65 ± 2.41

22.24 ± 2.77*

Retinol (mmol· L-1)

0.15 ± 0.20

0.28 ± 0.31

Lycopene (mmol· L-1)

0.03 ± 0.51

0.11 ± 0.10*

Beta carotene (mmol· L-1)

0.26 ± 0.65

0.07 ± 0.03

Alpha carotene (mmol· L-1)

0.55 ± 1.1

0.09 ± 0.08

Ascorbyl radical (Arbitrary units)

330893± 146373

418057 ± 174546

* Indicates a difference between group as a function of state (pre vs post scores) (P < 0.05).

7.2.4- One Repetition Maximum (1RM)- A significant relationship was observed for 1RM scores, (59.0 ± 8.74 vs. 67.31 ± 6.06, P<0.05). As can be seen in Table 7.2, 1RM scores increased by 14% following 8 weeks of antioxidant supplementation this trend was not expected.

7.2.5-Acute Resistance Scores- A significant interaction was demonstrated between pre and post acute resistance scores (54.30 ± 8.11 vs. 59.49 ± 8.59, P <0.05). As displayed in Table 7.2 8 weeks of antioxidant supplementation resulted in observed improvements of 9.5%.

7.2.6-Interleukin 6 (IL-6)- Following 8 weeks of antioxidant supplementation a significant relationship was observed for IL-6 (4.74 ± 1.10 vs. 3.43 ± 1.18, P <0.05). As displayed in Table 7.2, IL-6 decreased by 27% from baseline values.

7.2.7-Interleukin 10 (IL-10)- A significant relationship was established for IL-10 (4.39 ± 0.79 vs. 7.60 ± 1.88, P< 0.05). Following 8 weeks of antioxidant supplementation, IL-10 improved by 73% from pre chronic values.

7.2.8-Tumor Necrosis Factor α (TNFα)- No significant relationships were found for TNFα between pre and post supplementation (6.29 ± 0.41 vs. 6.55 ± 1.10, P>0.05). TNFα document marginal increases slightly following the 8 week supplementation period which was not expected.

7.2.9-Lipid Hydroperoxide (LOOH)- A significant relationship was established for LOOH (0.23 ± 0.14 vs. 1.55 ± 1.11, P<0.05). As demonstrated in Table 7.3 LOOH levels increased substantially following 8 week d of antioxidant supplementation.

Table 7.4- Body Composition analysis

Healthy Individuals

Pre Chronic

Post Chronic

Bone Mineral Density

1.05 ± 0.08

1.02 ± 0.08

T Score

-0.33± 0.75

-0.62 ± 0.82

Z Score

0.99 ± 0.74

0.79 ± 0.80

% Tissue Fat

38.23 ± 6.24

38.63 ± 6.37

% Region Fat

37.01 ± 6.61

37.48± 6.15

Centile

52.69± 25.20

54.42 ± 26.19

Total Mass

68.43 ± 8.21

68.11 ± 8.15

Tissue Mass

62435.62 ± 7752.85

62568.58 ± 7916.76

Fat Mass

24334.62 ± 6696.16

24572.58 ± 6900.53

Lean Mass

38100.92± 2662.05

37996.17 ± 2407.93

Bone Mineral Content

2092.69± 202.36

2062.67 ± 181.03

% Android Fat

41.6 ± 9.97

40.65 ± 11.83

% Gynoid Fat

47.9± 5.2

48.05 ± 5.21

A/G ratio

0.86 ± 0.15

0.84 ± 0.21

% Tissue Fat Mass

24.33± 6.70

24.57± 6.99

% Lean Mass

38.10 ± 2.66

38 ± 52.41

ALM/H2

5.96 ± 0.35

5.98 ± 0.35

* Indicates a difference between group as a function of state (pre vs post scores) (P < 0.05).

7.2.10-Gamma tocopherol- A significant relationship was observed for gamma tocopherol (1.5 ± 0.6 vs. 2.06 ± 0.44, P<0.05). 8 weeks of antioxidant supplementation increased levels by 37% from pre chronic values.

7.2.11-Alpha tocopherol- Following 8 weeks of antioxidant supplementation a significant relationship was observed for alpha tocopherol (19.65 ± 2.41 vs. 22.24 ± 2.77, P <0.05). As displayed in Table 7.3, Alpha tocopherol documented increases of 13% from baseline values.

7.2.12-Retinol- No significant relationships were found for retinol between pre and post supplementation (0.15 ± 0.20 vs. 0.28 ± 0.31, P>0.05). Although modest improvements were noted following 8 weeks of antioxidant supplementation.

7.2.13-Lycopene- Following 8 weeks of antioxidant supplementation a significant relationship was observed for alpha tocopherol (0.03 ± 0.51 vs. 0.11 ± 0.10, P <0.05). Lycopene concentration increased dramatically following 8 weeks of supplementation.

7.2.14-Beta carotene -No significant relationship was established for Beta carotene following 8 week supplementation phase (0.26 ± 0.65 vs. 0.07 ± 0.03, P> 0.05). However it must be noted that Beta carotene decreased by 73% from baseline values.

7.2.15-Alpha carotene- Following 8 weeks of antioxidant supplementation Alpha carotene decreased by over 80%, however this relationship was not found to be statistically meaningful (0.55 ± 1.1 vs. 0.09 ± 0.08, P>0.05).

7.2.16- Ascorbyl radical- As displayed in Table 7.3 Ascorbyl radical detection increased by 26% following 8 weeks of antioxidant supplementation, although this relationship did not elicit any significant relationship (330893± 146373 vs. 418057 ± 174546, P>0.05).

7.2.17- Body composition results as measured by DXA are presented in Table 7.4. As expected no significant differences were noted for bone mineral density, T score, lean mass, fat mass, total mass, fat percentage or bone mineral content (BMC) detected between treatments following 8 weeks of antioxidant supplementation.

7.3-Acute Resistance phase

7.3.1-Interleukin 6 (IL-6)- A significant relationship was observed for IL-6 following completion of acute resistance phases (P<0.05). As displayed in Figure 7.1 completion of acute resistance increased IL-6 expression, significant relationships were established between IL-6 at both acute resistance phases (pre acute 1 vs. post acute 1, P < 0.05) and (pre acute 2 vs. post acute 2, P < 0.05). Additionally significance was also established between (post 1 vs. post acute 2, P < 0.05).

Figure 7.1 displaying Interleukin 6 concentrations during all acute resistance exercise phases.

* Indicates a difference between group as a function of state (pre 1 vs. pre 2 scores) (P< 0.05).

7.3.2-Interleukin 10 (IL-10)- IL-10 demonstrated a significant relationship following completion of both acute resistance exercise phases (P<0.05). As can be seen in Figure 7.2 IL-10 was significantly higher during the pre acute 2 in comparison to pre acute 1 (pre acute 1 vs. pre acute 2, P < 0.05). Elevations were observed in IL-10 following completion of acute exercise during the second phase, (pre acute 2 vs. post acute 2, P <0.05).

Figure 7.2 displaying Interleukin 10 concentrations during all acute resistance exercise phases.

* Indicates a difference between group as a function of state (pre 1 vs. pre 2 scores) (P< 0.05).

7.3.3- Tumour Necrosis Factor α- A significant relationship was observed for TNFα following completion of both acute resistance exercise phases (P< 0.05). As displayed in Figure 7.3 TNFα significantly increased following completion of both the first (pre acute 1 v post acute 1, P < 0.05) and second acute resistance phase (pre acute 2 vs. post acute 2, P < 0.05). Additionally a meaningful relationship was also established between post acute phases (post acute 1 vs. post acute 2, P < 0.05), whereby TNF was notably higher during the post acute 2 timepoint.

Figure 3 displaying Tumour Necrosis Factor α concentrations during all acute resistance exercise phases.

* Indicates a difference between group as a function of state (pre 1 vs. pre 2 scores) (P< 0.05).

7.4- Discussion

The main finding of this study was that antioxidant supplementation over an 8 week period significantly improved the anti-inflammatory profile of healthy older females, documenting significant increases in IL-10 alongside reductions in IL-6. Additionally results demonstrate that 8 weeks of antioxidant supplementation, significantly increased alpha tocopherol, gamma tocopherol and lycopene, but significant increases in lipid hydroperoxides were also observed over the same time period, suggesting that antioxidant supplementation increased markers of oxidative stress and lipid peroxidation within the study cohort. Improvements were also observed for muscle strength measures over the 8 week time period, although the causative mechanism for this is not clear. However no improvements were noted in any markers of body composition over the 8 week time period as measured by iDXA. It is also important to note that completion of the chronic exercise period significantly altered the acute response to resistance exercise for IL-6, IL-10 and TNFα, which may be a positive finding in terms of the biochemical and functional status of skeletal muscle.

The current study documented significant increases in lipid hydroperoxide (0.23 ± 0.14 vs. 1.55 ± 1.11, P<0.05) following 8 weeks of antioxidant supplementation, whilst also reporting increased levels of both gamma and alpha tocopherol respectively (1.5 ± 0.6 vs. 2.06 ± 0.44, 19.65 ± 2.41 vs. 22.24 ± 2.77, P<0.05) . Lipid hydroperoxides are formed during the propagation phase in the process of lipid peroxidation (Packer et al., 2000). Research has suggested lipid hydroperoxide represent unstable molecules which can decompose to aledhydes or alternatively are rapidly broken down in cells through the actions of GSH peroxidise (Sen,Packer & Hänninen, 2000). Although both Vitamin E and Vitamin C have been shown to possess the ability to inhibit ROS initiated lipid peroxidation reactions (Chow & Chow-Johnson, 2013; Niki, 2013), this was not observed in the current study albeit with higher concentrations of both antioxidants constituents post supplementation phase. It has been suggested that Vitamin C and E are capable of exerting a protective effect through the reduction and prevention of oxidative damage (Packer et al.., 2000; Niki, 2013). Vitamin E has the ability to prevent lipid peroxidation chain reactions in cellular membranes by interfering with the propagation of lipid radicals (Chow & Chow-Johnson, 2013). Vitamin C predominately exerts its role as a reducing agent, reacting with Vitamin E radical to yield a vitamin C radical, resultantly regenerating Vitamin E (Packer et al., 2000). Considering the results in the current study the author would suggest that Vitamin C was able to effectively recycle Vitamin E, higher means ± SD following supplementation may be indicative of an active regenerative cycle between these antioxidants. Vitamin C (ascorbic acid) and Vitamin E (α-tocopherol) are frequently consumed in combination, it has been postulated that when co-ingested this permits an interaction between different antioxidant mechanisms obtaining a synergistic benefit (Packer et al., 2000; Petersen et al. 2001; Dawson et al., 2002; Fischer et al., 2004; Mastaloudis et al., 2004; Bloomer et al., 2006; Bloomer et al., 2007; Howatson et al., 2009). The antixodant supplement ingested in the current study also included lipoic acid. Packer et al., (1998) suggested that lipoic acid represented the missing link in terms of the antioxidant regeneration cycle, however results from the current study suggest that the addition of lipoic acid exerted a pro oxidant effect, potentially negating the scavenging and regenerative ability between tocopherol and ascorbyl. Research has demonstrated that higher levels of lipid hydroperoxides were reported in individuals who displayed signs of insulin resistance, whereby enhanced levels of insulin resistance were coupled with higher plasma concentration of lipid hydroperoxides (Nourooz-Zadeh, McCarthy S, Betteridge & Wolff, 1995; Facchini, Humphreys, DoNascimento, Abbasi & Reaven, 2000). Although this was examined in the current study, this may represent a plausible explanation for the higher not levels of lipid hydroperoxides. The authors suggested dietary intake played an important contributory role, postulating that reduced antioxidant intake could result in lower levels of carotenoids and tocopherols (Facchini, Humphreys, DoNascimento, Abbasi & Reaven, 2000). The net effect of which can potentially impair the ability of insulin to stimulate glucose disposal, conversely elevating levels of lipid peroxidation. Alternatively it was also proposed that lipid peroxidation, secondary to a decrease in antioxidant vitamins, might impair insulin action (Nourooz-Zadeh, McCarthy S, Betteridge & Wolff, 1995; Facchini, Humphreys, DoNascimento, Abbasi & Reaven, 2000).

A significant negative correlation was found between lipid hydroperoxides and ascorbyl radical levels in the present study (r= -0.409, P<0.05). As levels of lipid hydroperoxides increased, ascorbyl radical decreased. This is in agreement with research conducted by Vincent and colleagues, (2004) who demonstrated that Vitamin C intake was negatively correlated with lipid hydroperoxides (r=-0.707, P<0.05). Additionally the authors demonstrated that lipid hydroperoxides and TBARS levels were higher in obese individuals in comparison to non-obese individuals, although average BMI for the current study was (24.2 ± 3.11 ) (Vincent, Morgan, Vincent, 2004). Sabharwal & May, (2008) found that alpha lipoic acid in combination with Vitamin C exerted a protective effect on the endothelium and LDL from oxidative stress, however within this role they suggested that ALA did not spare ascorbic acid as part of this protection. Research has suggested that the recycling capabilities of tocopherol and ascorbyl may be heightened through the addition of lipoic acid through the reductioin of ALA to DHLA via NADH (Packer et al., 2000; Shay, Moreau, Smith & Hagen, 2006). Although current study findings do not offer support for this, the author suggests that lipoic acid in this instance may have exerted a pro oxidant effect. Lipoic acid has been highlighted in the research literature to be capable of combating ROS which may attenuate the oxidation of LDL-C, reducing arthrogenic and cardiovascular disease risk (Wollin & Jones, 2003), nonetheless evidence also exists documenting a pro-oxidant role (Niki, 2013). However as the current study protocol involves a supplementation phase only in the absence of exercise it is plausible that this may be a contributing factor towards the observed changes. Exercise has been shown to induce a protective effect against cellular oxidation through positive adaptations within the enzymatic antioxidant defence network (Ji, 2008). Both nuclear factor kappaB and mitogen-activated protein kinase signalling pathways have been implicated as causative factors to explain these changes, it is hoped that through examination of these transcription factors in the next chapter will provide answers in terms of the respective signalling pathway activated.

Currently debate exists as to whether LA exerts its function predominately as an anti-oxidation or pro-oxidant. This relationship is complex, whereby the resultant oxidant effect of LA is determined by the type of oxidant stress and is governed by the environmental stimuli (Gorca, Huk-Kolega, Piechota, Kleniewska & Ciejka, 2011). Packer et al., (2000) postulated that lipoic acid represented a highly effective therapeutic antioxidant, possessing the ability to be easily absorbed from the diet, easily converted into a usable form alongside the ability to interact with other important antioxidants such as ascorbic acid, a-tocopherol and glutathione in order to produce favour- able cardiovascular benefits (Packer et al, 2000; Shay et al., 2006; Pershadsingh, 2007; Peake, Katsuhik & Coombes, 2010). The current study is limited to the measurement of lipid hydroperoxides in terms of lipid peroxidation, it is plausible that lipoic acid negatively contributed towards the redox environment. The author suggests that in the absence of a measure of total antioxidant capacity, it is plausible that baseline levels of total antioxidant capacity may have been low, potentially explaining why no beneficial effect was observed for lipid hydroperoxides. Additionally in consideration of this fact, a longer duration may have been required in comparison to the 8 weeks currently completed in the study. Nevertheless, it seems reasonable an adequate antioxidant intake is needed to maintain healthy muscular activity (Maxwell 1995; Cerrulo et al., 2012). Research has proposed that inconsistent results may be attributable to targeting incorrect populations, whereby individuals should be chosen based on their potentially benefit such as those who have low antioxidant status or have experienced a clinical event such as surgery or fracture (Patrignani et al .,2000; Kris-Etherton, 2004; Cerrullo et al., 2012). This should be addressed in future research, in order to enhance knowledge into the relationship of dietary antioxidants intake with oxidative damage and serum antioxidants levels.

Together, results from the current study reinforce a complex relationship between oxidative stress and antioxidant supplementation. As displayed in Table 7.3 antioxidant supplementation increased levels of alpha tocopherol, gamma tocopherol, retinol, lycopene and ascorbyl. It is important to note that reduced levels of alpha and beta carotene and significantly higher levels of lipid hydroperoxides were also observed within the same frame. It has been suggested oral antioxidant supplementation can exert beneficial effects through a variety of different methods, it has been postulated that the predominate role of antioxidants is mediated via scavenging free radicals within different compartments (Fusco et al., 2007). Within the body’s first line of antioxidant defence, antioxidants work towards preventing the production of ROS/RNS through sequestering active metal ions and reducing peroxide concentrations (Fusco et al., 2007: Cerrulo et al., 2012). In their second defence line antioxidants scavenge, quench, or remove ROS/RNS and other reactive species before they attack biological molecules (Berger et al., 2005: Fusco et al., 2007). Following this antioxidants also possess the ability to repair the damage and reconstitute membranes and tissues (Diers, Landar, Darley-Usmar, 2012; Niki, 2013). It is important to note that recent research has suggested low levels of oxidative stress can induce an adaptive response, which can resultantly accelerate the production of antioxidant protein and enzymes (Niki, 2012). ROS have also been demonstrated to undertake an important signalling and regulatory role in certain circumstances (Ji, Gomez-Cabrera & Vina, 2009;Mc Ardle & Jackson, 2011). Resultantly the beneficial effect of antioxidant supplementation has been debated, posing the question does significantly higher level of antioxidants negate and disrupt the body’s natural response (Cerrulo et al., 2012). Summatively antioxidants exert their function in a synchronized and interrelated manner, to address higher levels of oxidative stress. The current study findings provide support for a synergistic link between Vitamin C and Vitamin E.

As documented in the results section alpha carotene (0.55 ± 1.1 vs. 0.09 ± 0.08, P >0.05) and beta carotene (0.26 ± 0.65 vs. 0.07 ± 0.03, P> 0.05) demonstrated marked decrements following 8 weeks of antioxidant supplementation. Research has highlighted that low serum/plasma carotenoids are independently associated with poor skeletal muscle strength and impaired physical performance in older individuals (Ceserai et al, 2004; Semba et al., 2007; Powers et al., 2010). The Women’s Health and Aging Studies (WHAS) I and II studied over 600 women aged 70-79 years old, demonstrating that low serum carotenoid levels were associated with poor muscle strength (Walston, Xue, Semba et al., 2006). However results from the current study offer alternate findings, although levels of alpha and beta carotene reduced during the supplementation period significant improvements were also noted in muscle strength during the same period. Research conducted by Young and colleagues, (2004) postulated that functionally carotenoids exert a role maintaining a balance with reactive oxygen species, findings from the study potentially offer support for this suggesting that they were attempting to address the redox environment. Serum carotenoids are believed to play an important role in reducing oxidative stress through the quenching of hydroxyl radicals and reducing lipid peroxidation (Mayne, 2003; Semba, Lauretani & Ferrucci, 2007). It is believed that via the reduction of free radical levels, carotenoids may positively modulate the cellular redox balance namely as a result of reduced activation of nuclear factor κB (NF-κB), a major transcriptional factor involved in the upregulation of interleukin-6 (IL-6) (Ershler & Keller, 2000; Kabe, Ando, Hirao, Yoshida & Handa, 2005; Semba, Bartali, Ferrucci, Ricks, Blaum & Fried, 2007). Morevoer, taking into consideration the significant increases in lipid hydroperoxides, it is plausible that reduced levels of carotenoids were an attempt to address the oxidative imbalance. Additionally in the InCHIANTI study, reduced levels of plasma carotenoids were independently linked to increased plasma IL-6 levels in elderly populations (Semba et al., 2003). The current study findings do not offer support for this, although observed decrements were noted in both alpha and beta carotene following the supplementation phase lower IL-6 was also reported as can be seen in Table 7.2. Nevertheless, it must be emphasised that such observations were noted over a longer period of time in comparison to the 8 week supplementation phase completed in the current study. On the other hand in consideration of the well documented relationship between heightened levels of IL-6 with enhanced risk of physical disability and sarcopenia, lower levels of IL-6 observed may be beneficial (Binder et al., 2004; Taylor et al., 2004). Diet represents an extremely important source of carotenoids, whereby the family of six major dietary carotenoids encompass a central component within the antioxidant defense system in humans protecting against oxidative stress by quenching singlet oxygen, scavenging free radicals, inhibiting lipid peroxidation, and modulating redox-sensitive transcription factors that are involved in the upregulation of proinflammatory cytokines (Hu et al., 2004; Semba, Lauretani & Ferrucci, 2007;Doria et al., 2012). As mentioned it may be plausible that changes occurred within the dietary patterns of these individuals, however as dietary assessment was not currently completed the author can only merely speculate.

The current study documented positive correlation between 1RM and alpha tocopherol (r=0.449, P<0.05). Currently a lack of consensus exists in the literature as to whether higher level of antioxidants can positively improve physical performance and muscular strength (Meng et al., 2010). Present study findings offer support for a role between 1RM and alpha tocopherol, which is in agreement with previous research within the area demonstrating improvements (Wijnen et al 2001; Takanami et al., 2000; Hauer et al., 2003; Upritchard et al., 2003; Gao et al., 2004). Interestingly research conducted by Jiang, Christen, Shigenaga, & Ames, (2001) and Ceseari et al., (2004) also demonstrated strong correlations for plasma antioxidant concentrations with physical performance and strength, specifically that of α-tocopherol. Lower levels of carotenoids and alpha-tocopherol have also been associated with lower grip, hip and knee strength (Semba et al., 2003), which has direct implications for the current study group, similarly the Hertfordshire Cohort Study demonstrated a positive association between beta-carotene and vitamin C with grip strength (Semba et al., 2003; Scott et al., 2010; Doria, Buonocore, Focarelli & Marzatico, 2012). However the current study findings adds debate to this issue, demonstrating increased ascorbyl radical production alongside reduced alpha and beta carotene, but participants experienced improvements in hand grip strength scores. Ryan and colleagues, (2008) demonstrated in animal models that an antioxidant supplemented diet maintained force production in comparison to a control diet, resultantly eliciting an improved ability to produce positive work when supplemented with Vitamin E and C. The authors proposed that together Vitamin C and E positively improved the redox environment, resulting in lower levels of basal oxidative stress. It is plausible in the current study enhanced levels of antioxidants post supplementation may have exerted a beneficial effect on force production. Sarcopenia as a condition has many clinical manifestations, the age associated loss of muscle mass and strength is a focal contributor towards physical disability and functional impairment (Morley et al., 2001; Taylor et al., 2004). Accumulative evidence supports the role of increased oxidative stress in the progression of sarcopenia (Hianna, Jan, Douglas, Stuart & Tarnoplosky, 2004; Meng et al., 2010). Although research has documented higher levels of oxidative damage to DNA, protein, and lipids with advancing age, the question remains as to whether oxidative stress represents an initiating event in the progression of sarcopenia or a compensatory reaction is inconclusive (Semba, Bartali, Ferrucci, Ricks, Blaum & Fried, 2007; Doria, Buonocore, Focarelli & Marzatico, 2012). Hence, the relationship between sarcopenia and dietary antioxidants is not yet fully elucidated. In the current study improvements in α-tocopherol and γ- tocopherol were accompanied by significant improvements in lower extremity leg strength suggesting a link between heightened levels of tocopherol and improvements in leg strength.

7.5-Conclusion.

Lack of dietary assessment presents a major drawback for the current study, which may have potentially offered explanations towards a number of observed changes. The current study protocol involved a supplementation phase only in the absence of exercise, the author suggests that perhaps supplementation with exercise may have been more efficacious. Delays were experienced in the production of the antioxidant supplementations, due to shortage in vitamin E this changed our target population ideally we would have liked to supplemented post hip fracture females. However due to the time constraints of the current project this was not feasible. Additionally the author suggests that a measure of total antioxidant capacity would have been a beneficial inclusion. Summatively the current study adds further debate to the conflicting research findings for antioxidant supplementation within elderly populations. Due to the significant increases in lipid hydroperoxides, the author suggests that lipoic acid may have exerted a pro-oxidant role, even though increases in tocopherol and ascorbyl radical were noted. Current study findings demonstrate a complex relationship between oxidative stress and antioxidant supplementation, however in light of observed findings the author would not support the ingestion of the prescribed antioxidant cocktail containing Alpha lipoic acid, Vitamin C and Vitamin E. However it is hoped that the application of DNA microarray work, will provide answers into transcriptional activation and signalling pathways activated pre and post antioxidant supplementation which may reveal avenues for future research.