Veränderungen des Skelettmuskels bei Tumorkachexie Morphologische, metabolische und Genexpressions-Analyse des humanen Musculus rectus abdominis
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Philipps-Universität Marburg
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Abstract
The complex syndrome of cancer-associated cachexia, an unintentional loss of muscle (and fat)
mass, presents an oncological challenge: It reduces the patients’ quality of life and mobility and
may lead to death e.g. via respiratory muscle insufficiency. 20% of all carcinoma patients are
estimated to die of cancer cachexia, yet neither is the underlying mechanism fully understood
nor is the condition therapeutically controllable. Studies on human tissue samples, in particular
on skeletal muscle, are scarce and have yielded ambiguous results in contrast to those in cell
cultures and animal models. Pancreatic carcinoma is the clinically most impressive example of a
malignancy with the most severe and rapid cachexia development. Therefore the aim of this
thesis was to evaluate the cancer cachexia-associated changes in skeletal muscle biopsies from
patients with pancreatic cancer at the time of their first surgery. The analyses comprised muscle
fiber morphometry, the concentration of metabolically important amino acids and the
expression of pro-inflammatory, -angiogenetic, -atrophic and -apoptotic signals on RNA as well
as protein levels. The study included intraoperative skeletal muscle biopsies from the rectus
abdominis muscle of 36 patients with histologically confirmed ductal pancreatic carcinoma. The
group of patients with cachexia (N=16) was compared with that without cachexia (N=22, control
group). Cancer cachexia was defined as unintentional weight loss of at least 10% over the past
6 months. Transverse cryosections of rectus abdominis muscle were stained for myofibrillar
ATPase to measure size, density and composition of muscle fibers, for CD31 to measure capillary
contacts and for haemalum to measure centralized nuclei. In addition, the intracellular
concentration of 26 amino acids was analyzed by HPLC, differential expression of 29 genes was
determined via qRT-PCR, and pools of tissue samples were screened for 102 proteins using a
protein array.
Contrary to expectations, no significant decrease in muscle fiber size or increase in density as in
muscle atrophy was observed with cachexia compared to the control group. Neither cachexiaassociated
upregulation of muscle-specific E3-ligases TRIM63 and FBXO32 of the ubiquitin
system, commonly used as proteolysis markers, nor upregulation of BAX and CASP3 (proapoptotic
signals) were detected. But the protein expression of Fas ligand was 2.32-fold
increased and the expression of RNA of the antiapoptotic BCL2 was significantly 0,49-fold lower
(p=0.028). Abundance of cells with centralized nuclei was significantly increased by 7% (p=0.031)
in muscle fiber type I and over all fiber types by 5% (p=0.085), likely indicating increased muscle
regeneration, though PAX7 expression remained unaltered. As a main new finding, there was a
significant 20% (p=0.022) decrease in the intracellular concentration of the proteinanabolic and
antiproteolytic amino acid leucine, which, notably, correlated to weight loss (p=0.046). The
concomitant finding of 1.782- and 1.822-fold increased expression of the transmembrane
transporters SNAT2 (p=0.013) and LAT1 (p=0.072), respectively, did not allow any conclusion on
(limited) leucine uptake, especially as no blood samples were available for extracellular leucine
measurements.
The present study provided evidence for a pro-inflammatory milieu with cachexia compared to
controls in terms of a significantly increased expression of CD68 (1.48-fold, p=0.040), indicating
macrophage infiltration, and, moreover, a significantly decreased relative expression of GCS
(0.831-fold; p=0.036) and GSR (0.657-fold, p=0.026) as involved in syntheses and reduction of
the antioxidant glutathione, respectively. Correspondingly the following inflammatory factors
were increased in the protein array: IL1β (1.41-fold), IL6 (1.50-fold), IL8 (1.48-fold), TNF (1.87-
fold), interferon-y (1.14x), CRP (1.55-fold), and IL32 (3.03-fold). The mRNA expressions of IL1β,
IL6, IL8 and TNF were, however, not significantly increased. The mRNA expressions of the
monoamine oxidases, as a possible source of oxidative stress, were inconsistent, as changes in
MAOA expression were 0.19-fold (p=0.030) while those of MAOB were 1.4-fold (p=0.047). In line
with pro-inflammatory changes, a 33% increase in capillary contacts of the muscle fibers
(p=0.097) was observed along with, by trend, higher protein levels of pro-angiogenetic signals:
VEGF was 1.30-fold, angipoietin-1 2.07-fold, Angiopoietin-2 1.77-fold increased and PF4 was
0.16-fold suppressed. The mRNA expressions of VEGFA, VEGFB and KDR were, however,
unchanged.
In synopsis of these new human data on human skeletal muscle from patients with pancreatic
cancer, it can be stated that at an early pre-atrophic stage of cachexia intracellular leucine, as
an essential anabolic mediator, is significantly reduced. At the same time there is an
inflammatory milieu associated with a trend towards angioneogenesis and possibly increased
muscle regeneration. This early metabolic and inflammatory condition in not yet atrophied
skeletal muscle may be relevant as a possible target for prevention and a therapeutic approach
to cancer cachexia. A relevant time window and a set of parameters for characterization of a
“myositis-like-phase” of cachexia development were identified for further clinical studies.
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Dates
Created: 2020Issued: 2020-12-07Updated: 2020-12-07
Faculty
Medizin
Publisher
Philipps-Universität Marburg
Language
ger
Data types
DoctoralThesis
Keywords
skeletal musclePankreaskarzinom;KachexieSkelettmuskelcancer cachexia
DFG-subjects
SkelettmuskelKachexiePankreaskarzinom;
DDC-Numbers
610
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Rockenbach, Johanna Serena: Veränderungen des Skelettmuskels bei Tumorkachexie Morphologische, metabolische und Genexpressions-Analyse des humanen Musculus rectus abdominis. : Philipps-Universität Marburg 2020-12-07. DOI: https://doi.org/10.17192/z2020.0260.