Introduction
Diabetic kidney disease (DKD) is a prevalent metabolic disease and is
the most common cause of end stage renal disease (ESRD) (Fu et al.,
2019). Despite existing therapies that target hypertension and
hyperglycemia, these treatments only slow the progression to ESRD. Thus,
there is a need for the identification and development of new
pharmacological therapeutics to treat DKD.
In type 2 diabetes, early hyperglycemia and glomerular hyperfiltration
affect renal glomerular and proximal tubular function. Hyperglycemia
exposes the proximal tubule to increased amounts of filtered glucose,
which in turn leads to increased glucose reabsorption (Vallon, 2011).
The increased tubular glucose load results in a number of
pathophysiological changes: proximal tubule growth, upregulation of
sodium-glucose cotransporter 2 (SGLT2), inflammation, mitochondrial
dysfunction and eventually leading to tubulointerstitial fibrosis
(Abbate & Remuzzi, 1999; Bohle et al., 1991; Cleveland et al., 2020;
Forbes & Thorburn, 2018; Huang & Preisig, 2000; Vallon & Verma,
2021). Although there are clear and distinct changes in proximal tubular
function in DKD, the mechanisms underlying these changes are
understudied.
The kidney is a highly metabolic organ that relies heavily on
mitochondrial oxygen consumption to account for the energy requirements
of tubular reabsorption (Bhargava & Schnellmann, 2017; Lynch et al.,
2018). Mitochondrial dysfunction has been identified as a key event in
the early stages of hyperglycemia leading to disease progression. Renal
mitochondrial dysfunction encompasses multiple functional changes.
Studies have shown that expression and activity of the master regulator
of mitochondrial biogenesis (MB) peroxisome proliferator-activated
receptor gamma co-activator 1 alpha (PGC1α) is downregulated in proximal
tubules of diabetic animals, leading to disease progression (Lee et al.,
2017).
In addition, studies report that mitochondrial dynamics is altered in
diabetic db/db mice and in glomerular podocytes exposed to high glucose
(Ayanga et al., 2016; Galvan et al., 2019). These changes in
mitochondrial dynamics were mediated by the mitochondrial fission
protein dynamin-like protein 1 (Drp1). When Drp1 was knocked down or
pharmacologically inhibited, mitochondrial function was restored.
Importantly, restoration of mitochondrial dynamics and function further
led to significant improvement of hallmark features of DKD including
glomerular scarring, albuminuria and mesangial matrix expansion. It has
also been reported that Mfn1 expression decreases in response to high
glucose, although the extent of knowledge regarding Mfn1 in DKD is
limited (Audzeyenka et al., 2021). These studies provide evidence that
mitochondrial function is altered in response to glucose and improving
aspects of mitochondrial dysfunction such as mitochondrial dynamics can
be an effective therapeutic strategy in DKD.
We previously demonstrated that in RPTC exposed to high glucose,
phosphorylation of the mitochondrial fission protein Drp1 was increased
and expression the mitochondrial fusion protein Mfn1 was decreased,
indicating that there is an imbalance in RPTC mitochondrial dynamics in
the presence of glucose (Cleveland et al., 2020). Interestingly, the
same effects were observed in renal cortical tissue of early diabetic
db/db mice. Despite these clear and distinct alterations in
mitochondrial dynamics proteins, the signaling mechanisms underlying
these effects remain unknown. Studies have identified that Drp1
phosphorylation in hyperglycemia is mediated by Rho-associated protein
kinase 1 (ROCK1) (W. Wang et al., 2012). Furthermore, separate studies
show that Ras homolog family member A (RhoA) is responsible for ROCK1
activation and subsequent Drp1 phosphorylation (Brand et al., 2018).
Little is known about the role of Mfn1 and its associated signaling
pathways in high glucose. However, it has been shown that MEK1/2/ERK1/2
regulates Mfn1 function (Pyakurel et al., 2015). In addition, it has
been demonstrated that Raf, the upstream activator of MEK1/2 is also
upregulated in the presence of glucose (Trumper et al., 2005).
Our previous study showed that altered Drp1 phosphorylation and Mfn1
expression in diabetic db/db mice as well as in RPTC exposed to high
glucose was restored by treatment with the
β2-adrenoceptor agonist formoterol. However, the
mechanisms as to how formoterol activation of the
β2-adrenoceptor regulates mitochondrial dynamics have
yet to be determined. In separate studies, we also demonstrated that
formoterol promotes recovery from acute kidney injury (AKI) by
stimulating MB (Jesinkey et al., 2014; Wills et al., 2012). Despite the
well-known classical pathway of β2-adrenoceptor
activation, defined by Gαs stimulation of adenylyl cyclase (AC) and cAMP
production, we discovered that formoterol signals through Gβγ to
activate Akt/eNOS/sGC/PGC1α to induce MB (Cameron et al., 2017). Based
on these findings, our hypothesis was formoterol signals through the Gβγ
subunit of the β2-adrenoceptor to regulate
RhoA/ROCK1/Drp1 and Raf/MEK1/2/ERK1/2/Mfn1 to restore the imbalance
between mitochondrial fission and fusion in DKD.