Mitochondrial health and function.

Mitochondrial health is vitally important for cellular viability and function (Magal- haes, Venditti et al. 2014). As the powerhouse of the cell, mitochondria are responsi- ble for liberation of the energetic substrate, ATP via the fatty acid cycle and oxidative

phosphorylation (Figure 1.8)  (Hafner, Brown et al. 1990).The symbiotic relationship

between the mitochondria and nucleus enables nuclear synthesis of the majority of the 1500 mitochondrial proteins (37 are encoded in the mitochondrial genome, 13 of which are required for oxidative phosphorylation) (Anderson, Bankier et al. 1981, Gaston, Tsaousis et al. 2009).These include subunit 6 (mt-ND6) of NADH dehydro- genase (complex I) and subunit 6 (mt-ATP6) of ATP synthase (complex V) (Van Ber-

gen, Blake et al. 2014). Given the critical role of mitochondria  to cellular health, the

process of nuclear synthesis of mitochondrial proteins may have arisen to minimise the effect of the greater mitochondrial mutation rate, as much as 10-20 times that of the nuclear genome (De Pauw, Demine et al. 2012). Coupled with this, mitochondrial DNA repair enzymes  have a low activity status (Linnane, Marzuki et al. 1989, Wang, Lin et al. 2009). Therefore, quality control mechanisms have evolved within the mito- chondria in an attempt to overcome some of the limitations of this inefficient DNA repair system. One example of these quality control mechanisms is mitochondrial biogenesis (Attardi and Schatz 1988, Medeiros 2008, Ren, Pulakat et al. 2010, Weck- becker and Herrmann 2013).

1.3.1. Mitochondrial biogenesis.

Mitochondrial biogenesis protects mitochondrial health through the maintenance of mtDNA quality (Scarpulla, Vega et al. 2012). This process is achieved via cycles in-

volving the coalescence of old mitochondria and their subsequent division (Ren, Pu- lakat et al. 2010). As a result, functional mitochondrial DNA is retained whilst com- promised mitochondrial DNA is selectively eliminated (Ren, Pulakat et al. 2010). Mi-

tochondrial DNA synthesis occurs de novo during the cell cycle to return mitochon-

dria mtDNA abundance to their homeostatic levels (Sheng and Cai 2012). However, during obesogenic conditions, mitochondrial biogenesis is impaired thereby contribut- ing to mitochondrial dysfunction. Mitochondrial biogenesis is coordinated at a tran- scriptional level by peroxisome proliferator activated receptor co-activator 1 alpha (PGC1α) (Fernandez-Marcos and Auwerx 2011) and nuclear respiratory factor 1 (NRF-1) (Escriva, Rodriguez-Pena et al. 1999). Together these promote the induction of the catalytic subunit of the mitochondrial DNA polymerase (POLG) and expression of other biogenesis transcription factors i.e. mitochondrial transcription factor A (Tfam) and oxidative phosphorylation (OXPHOS) genes required for mitochondrial replication (Murholm, Dixen et al. 2009, Elachouri, Vidoni et al. 2011). Previous studies have identified that patients who possess mutations in either a mitochondrial or nuclear gene involved in mitochondrial function, exhibit disturbances in their respi-

ratory capacity (Pich, Bach et al. 2005). Thus  respiratory or oxidative capacity is a

reflection of both mitochondrial biogenesis and mitochondrial morphology regulated via mitochondrial dynamics.

1.3.2. Mitochondrial dysfunction.

Mitochondrial dysfunction is most commonly defined as the inability of mitochondria to produce ATP in response to cellular demand (Bournat and Brown 2010). Neverthe- less, this definition can be extended to incorporate changes to mitochondrial biogene-

sis, reduction in fatty acid oxidation or increased production of reactive oxygen spe- cies (ROS) (Lowell and Shulman 2005, Zorzano, Liesa et al. 2009).

Figure 1.5. Origins of mitochondrial dysfunction.

Mitochondrial quality control involves surveillance and protection strategies to limit mitochondrial damage and ensure mitochondrial integrity. This quality control occurs at the following molecular, organelle and cellular levels. Figure and legend taken

from (Sheng and Cai 2012).

Importantly, mitochondrial dysfunction has been proposed to account for the genera- tion of toxic lipid metabolites via the fatty acid cycle and hampering of metabolic fit- ness, which accompany the stunted adipogenic capacity and Adipose tissue dysfunc-

tion (Vankoningsloo, Piens et al. 2005, Liu, Lin et al. 2012). Finally,  evidence in fa-

co-association of reduced mitochondrial oxidative phosphorylation capacity and insu- lin resistance in elderly or obese individuals (Petersen, Dufour et al. 2004). Further- more, mitochondrial dysfunction extends beyond the inability to synthesise sufficient ATP, to processes of mitochondrial DNA abundance and mitochondrial morphology (Okamoto and Shaw 2005, Yu, Robotham et al. 2006). In the skeletal muscle of obese and T2DM, a reduction of mitochondrial copy number together with enhanced mito- chondrial fission have been observed (Barthelemy, Ogier de Baulny et al. 2001, Jeng, Yeh et al. 2008). Mitochondrial dynamics, which involves mitochondria fission, is discussed in further detail in section 1.4.2.

1.3.3. Causes of mitochondrial dysfunction.

Despite multiple studies in skeletal muscle implicating a connection between insulin resistance and mitochondrial dysfunction, little data is available in white adipose tis- sue (WAT), regarding the involvement of mitochondria during conditions of insulin resistance or AT dysfunction.

1.3.4. Mitochondrial dysfunction in adipose tissue.

In humans, insulin resistance has been found to co-exist with a lower mitochondrial oxidative phosphorylation activity (Petersen, Dufour et al. 2004). Nevertheless, whether insulin resistance is a primary or secondary effect of mitochondrial dysfunc- tion is unknown (Shulman 1999, Shulman 2000, Guilherme, Virbasius et al. 2008, Amati, Dube et al. 2011, Koliaki and Roden 2014). In an attempt to address this ques- tion, studies performed in skeletal muscle of obese and T2DM subjects found that these cells are often exposed to a high fat load (Sparks, Xie et al. 2005, Catta-Preta,

Martins et al. 2012). Recapitulating this scenario and analysis of mitochondrial dy- namics in skeletal muscle biopsies from either morbidly obese (Semple, Crowley et al. 2004), following a high fat feed in humans with an ideal BMI (Richardson,

Kashyap et al. 2005, Sparks, Xie et al. 2005)or mice (Rong, Qiu et al. 2007) resulted

in the downregulation of the mitochondrial biogenesis gene, PGC1α (Puigserver and Spiegelman 2003, Mitra, Nogee et al. 2012). As mentioned earlier in section 1.3.1, the respiratory capacity of mitochondria is a reflection of both mitochondrial biogenesis and morphology that are regulated via mitochondrial dynamics.