Muscle Atrophy in Lung Cancer and Hemodialysed Patients Clinical Trial
Official title:
Study of Muscle Wasting Mechanisms and Biomarkers in Hemodialysis Patient
Muscle wasting is present in almost 50% of patients treated with chronic hemodialysis. It is
associated with an increased risk of death (particularly from cardiovascular causes) and
compromises quality of life (loss of autonomy and fatigue). The mechanisms leading to muscle
wasting in chronic kidney disease have been the subject of several studies in animals. These
have highlighted the role of the ubiquitin-proteasome system (UPS). Activation of UPS during
chronic kidney disease is multifactorial. It is the result of resistance to the action of
insulin/IGF1, metabolic acidosis, low grade prolonged inflammation and increased production
of myostatin. To date few studies have been conducted in humans.
The investigators want to identify blood markers related to muscle protein breakdown in
patients undergoing hemodialysis. In parallel, the investigators want to adress the
mechanisms involved in muscle proteolysis. In addition, the investigators want to identify
the proteins degraded and the ubiquitination enzymes (E2/E3 couples) specifically involved in
muscle loss during hemodialysis.
Muscle biopsies and blood sample will beperformed during scheduled surgeries in healthy
volunteers (negative control), cancer patients (positive control) or undergoing chronic
hemodialysis. RNA seq analysis will be performed in blood samples and proteomic mass
spectrometry analysis for establishing a specific profile between muscle and blood markers. A
limited subset of blood markers common to cancer and hemodialysis atrophying muscles will be
used for elaborating a chip dedicated to early detect an atrophying process.
Thus, the investigators will first design a diagnostic tool for detecting non-invasively
muscle protein breakdown before the onset of muscle atrophy. This will enable early and
efficient nutritional counter-measures.
Hypothesis Measuring urinary 3-methylhistidine is a simple tool to assess proteolysis in
clinical practice. However it is not totally specific of muscle proteolysis because it may
reflect an intestinal catabolism. Moreover, its dosage cannot be used in patients with renal
impairment especially in case of anuria. Apart from achieving muscle biopsy or measuring the
overall kinetics of proteins it does not yet exist simple and reproducible diagnostic tool
for measuring muscle proteolysis. With the present study, the investigators expect to
identify in the blood of patients a particular transcriptomic profile related to the
activation of muscle proteolysis, including increased proinflammatory cytokine transcripts.
Rodent models of CKD were used to identify major proteolytic system and signal pathway
implicated in muscle wasting caused by uremia. These finding needs to be confirmed in humans
studies. Also, other actors of muscle atrophy could be identified in humans. Notably, so far,
few studies have been performed in order to determine which E2 are involved in muscle
atrophy, and to knowledge of the investigators no one in CKD. Finally, data on muscle
proteasome substrates are limited since only actin was clearly identified in humans by the
team. The investigators expect an activation of the proteasome system with an increase in the
expression of enzymes E2s, E3s including MURF1 and MAFbx and proteasome subunit in patients
treated with hemodialysis. The investigators could also observe an activation of caspase
system which is thought to work in concert with the UPS.
Materials and methods 2.1. Population The experiments will be performed using blood sample
and human muscle biopsies obtained during programmed orthopedic surgery (negative control),
cancer (positive control) and hemodialysis patients. This translational research program has
already been accepted (DGS 2008-A00479-46) and inclusion of patients already started. A
maximum of 15 patients per group will be included. Inclusion and exclusion criteria were
set-up according to the objectives (absence of any pathology for controls, etc.) and the
legislation (written consent, etc.). Thirty-two patients have already been included so far,
and biopsies are stored in liquid nitrogen. Blood samples are collected just before surgery
using Paxgene RNA tubes and stored following the manufacturer's instructions until analysis.
2.2. Blood sample Total RNA will be extracted from blood using Paxgene RNA Extraction kit and
analysed by RNAseq 2.3. Muscle biopsies 2.3.1. Transcriptomic analysis Total RNA will be
extracted from human muscle biopsies using commercial kits. The investigators will first
verify that known muscles markers are up-regulated in cancer patient biopsies (positive
controls) and address their levels in muscle biopsies from hemodialysis patients. We will
thus perform qRT-PCR UBE2B, UBE2D2, MaFbx, MuRF1, Ozz, ASB2, E4B, Nedd4, Mdm2 and 26S
proteasome subunits.
2.3.2. Proteomic analysis Proteomic analysis will be performed usin shot gun spectrometry
analysis
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