The ability of the growth hormone secretagogue (GHS) Ipamorelin to counteract the catabolic effects of glucocorticoid (GC) on skeletal muscles and bone was investigated in vivo in an adult rat model. Groups of 8-month-old female rats were injected subcutaneously for 3 months with GC (methylprednisolone) 9 mg/kg/day or GHS (Ipamorelin) 100 microg/kg three times daily, or both GC and GHS in combination. The maximum tetanic tension of the calf muscles was determined in vivo in a materials testing machine. The maximum tetanic tension was increased significantly, and the periosteal bone formation rate increased four-fold in animals injected with GC and GHS in combination, compared with the group injected with GC alone. In conclusion, the decrease in muscle strength and bone formation found in GC-injected rats was counteracted by simultaneous administration of the growth hormone secretagogue.
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Growth Hormone & IGF Research 2001, 11, 266–272
doi:10.1054/ghir.2001.0239, available online at http://www.idealibrary.com on
The growth hormone secretagogue
ipamorelin counteracts
glucocorticoid-induced decrease
in bone formation of adult rats
N. B. Andersen1, K. Malml ̈of2, P. B. Johansen2, T. T. Andreassen1, G. Ørtoft1
and H. Oxlund1
1Department of Connective Tissue Biology, Institute of Anatomy, University of Aarhus, DK-8000 Aarhus C, Denmark and 2Novo Nordisk A/S,
DK-2760 Maaloev, Denmark
Summary The ability of the growth hormone secretagogue (GHS) Ipamorelin to counteract the catabolic effects
of glucocorticoid (GC) on skeletal muscles and bone was investigated in vivo in an adult rat model. Groups of
8-month-old female rats were injected subcutaneously for 3 months with GC (methylprednisolone) 9 mg/kg/day
or GHS (Ipamorelin) 100 μg/kg three times daily, or both GC and GHS in combination. The maximum tetanic
tension of the calf muscles was determined in vivo in a materials testing machine. The maximum tetanic tension
was increased significantly, and the periosteal bone formation rate increased four-fold in animals injected with
GC and GHS in combination, compared with the group injected with GC alone. In conclusion, the decrease in
muscle strength and bone formation found in GC-injected rats was counteracted by simultaneous administration
of the growth hormone secretagogue.
C°2001 Harcourt Publishers Ltd
Key words: growth hormone secretagogue, muscle tetanic tension, bone formation, bone bending strength,
muscle strength, glucocorticoid.
INTRODUCTION
Long-term glucocorticoid treatment may result in
weakening of both skeletal musculature1,2, 3 and bone
structures4,5 in man. The interaction between gluco-
Received 19 March 2001
Revised 12 July 2001
Accepted 16 July 2001
Correspondence to: Hans Oxlund, MD, DMSc, Department of Connective
Tissue Biology, Institute of Anatomy, University of Aarhus, DK-8000 Aarhus
C, Denmark, Tel: +45 89423017; E-mail: [email protected]
The data were presented at the 3rd International Conference of the Growth
Hormone Society, San Francisco, CA, 1998: Andersen NB, Oxlund H,
Malml ̈of K, Johansen PB, Ørtoft G, Andreassen TT. Effects of Ipamorelin—a
growth hormone secretagogue—on bone formation and skeletal muscle
contraction force. Growth Horm IGF Res 1998; 8: 313 (abstract).
corticoid-induced muscle weakness and osteopo-
rosis may lead to serious disability. Frail patients would
certainly benefit from a treatment regimen which could
increase skeletal muscle and bone strength.
We have developed an experimental animal model6,7
to study the ability of a growth hormone secretagogue
(GHS) to counteract glucocorticoid (GC) effects on
skeletal musculature and bone. In animal experiments,
a decrease in muscle protein synthesis,8muscle mass
and muscle strength has been observed in GC-treated
rats,9,10 as well as a reduction in bone mass and
strength.11,12
Growth hormone (GH) administration increases
muscle fibre size,13,14 leading to increased skeletal
muscle mass,15 which is accompanied by a propor-
tionate increase in muscle strength.7Furthermore, GH
1096–6374/01/050266+07 $35.00/0 C°2001 Harcourt Publishers Ltd
GHS counteracts GC-induced effects on bone 267
stimulates bone formation and increases bone
mass.15,16, 17 Growth hormone secretagogue may also
increase the bone mineral content of young rats18
and promote muscle function. Lieber et al.19 reported
an increase in muscle strength and fibre size in dogs
treated with GHS during remobilization.
Growth hormone administration may counteract the
GC-induced catabolic effects on muscle and bone
mass.11,20 GHS may reverse GC suppression of GH
secretion.21,22 At present it is not known, however,
to what extent GHS can counteract the catabolic ef-
fects of GC. The GHS, Ipamorelin, is a pentapeptide
selective for GH release similar to that displayed by
growth hormone releasing hormone.23 In short-term
studies, Ipamorelin increases BW gain, in GC-treated
animals24 and increases longitudinal growth in young
rats.25
The effects of the GHS, Ipamorelin, and GC admin-
istration, alone and in combination, were investigated
on muscle strength, muscle mass, bone mass, bone for-
mation rate and bone strength.
MATERIALS AND METHODS
Seventy female Wistar Hannover 8-month-old rats,
from Møllegaard Breeding and Research Centre
(Ll. Skensved, Denmark) were randomly divided into
five groups with 14 rats in each group. Group A
was killed at the start of the experiment. Group B
was injected daily with saline. Group C was injected
with GC 9 mg/kg/day (Solumedrol, Upjohn, Puurs,
Belgium). Group D was injected with GHS, 100 μg/kg
(Ipamorelin, Novo Nordisk A/S, Gentofte, Denmark)
three times daily. Group E was injected with a combi-
nation of GC and GHS in the above mentioned doses.
Injections were given subcutaneously in the nape of
the neck daily for 3 months. The animals had free ac-
cess to drinking water and normal pellet diet (Altromin,
Chr. Pedersen, Ringsted, Denmark), in a 12 hours light,
12 hours of darkness cycle. All rats were injected subcu-
taneously with 20 mg tetracycline/kg (Sigma Chemical
Co., St. Louis, MO, USA) 25 days before killing, and with
15 mg/kg of calcein (Sigma Chemical Co., St. Louis, MO,
USA) 5 days before killing. The study was approved by
the Danish Animal Experiment Inspectorate.
Maximum tetanic tension of skeletal muscles
The maximal tetanic tension (MTT) and time course of
tetanic tension of the calf musculature (m. soleus, m.
plantaris and m. gastrocnemius together) were studied
in vivo using a previously described method.7The rats
were anesthetized with pentobarbital 60 mg/kg in-
traperitoneally. The proximal end of the three mus-
cles was kept in position by a 0.7 mm steel wire
through a 1 mm transverse canal drilled through the
centre of the femur condyles. The distal tendons of
the muscles were fixed with a pin to maintain nat-
ural in situ length, and the calcaneus was then cut
and the distal end of the tendons was clamped and
fixed to the movable cross-head of a materials test-
ing machine. The femur epicondyles were fixed by
the steel wire to the load cell of the materials test-
ing machine (TCT5, Lorentzen & Wettre, Stokholm,
Sweden). The muscle tension was measured continu-
ously by means of the load cell, the direct signals fed
to a load-time recorder (Philips PM 8272, Eindhoven,
Netherlands).
The muscles were stimulated to contraction through
n. ischiadicus on the back of the thigh. The elec-
trical stimulus was given through steel electrodes
(diameter 1.2 mm, electrode distance 1 mm) which
were connected to a stimulator (Anapulse stimulator,
model 302-T, W-P Instruments, New Haven, CO, USA).
The electrodes were isolated from the surrounding tis-
sue with parafilm to avoid activation of adjacent mus-
cles and nerves. During the experiment the rats were
placed on a small electric blanket to maintain constant
body temperature.
At the beginning of the experiment, the muscles
were strained until a tension of 4 N was obtained.
The first two stimulations were single stimulations
(30 V for 4 ms) and were given to ensure a correct local-
ization of the electrodes. Then two stimulations of 1-s
duration (30 V, impulse-duration 4 ms, a frequency of
100 Hz) were given to induce maximal tetanic contrac-
tion force. The final stimulation was 30 s in duration
(30 V, impulse-duration 4 ms, a frequency of 100 Hz).
Maximal tetanic tension of each rat was calculated as
the mean value of the maximal tetanic tension obtained
in the last three contractions. The endurance of the
muscles was studied by following the time course of
the tetanic tension during 30 s stimulation. The rats
were killed by exsanguination, under anaesthesia. The
blood was centrifuged and serum was stored at −20◦C
until analysis.
Skeletal muscle mass
The hind legs were exarticulated in the hip joints. The
calf muscles were dissected free from the hind legs,
defatted for 7 days in three changes of acetone, freeze-
dried for 3 days and weighed (dry defatted weight =
DDW). The tibia bone length was measured from facies
articularis condylus medialis to facies articularis infe-
rior with a sliding caliper.
268 N.B. Andersen et al.
Mechanical three-point bending analysis
of cortical bone
The Strength of the femur diaphyses were assessed in
a materials testing machine (Alwetron 250, Lorentzen
& Wettre, Stockholm, Sweden), using a standardized
procedure,15,17, 26 The femurs were placed on the pos-
terior surface on two rounded supporting bars with a
distance of 15.1 mm between the bars. A third rounded
bar was then lowered onto the anterior surface, load-
ing the bone diaphysis. The loading point was at a dis-
tance of 55% of the total length of the femur from the
top of the caput femoris. The femur was deflected at
a constant speed of 5 mm/min until failure and load-
deflection curves were registered simultaneously on
an x-y-recorder by transducers coupled to measuring
bridges. The ultimate breaking load (bending strength)
was calculated.
Bone formation and histomorphometry
Two 300 μm transverse sections were cut from both
femurs 2 mm below the loading point by means
of a precision bone saw (Eksakt-Apparatebau, Otto
Hermann, Norderstedt, Germany). One of the sections
was placed on a slide and projected onto a screen by
means of a projection microscope, magnification 20x.
The outer and inner circumferences were drawn and
scanned into a computer. Circumferences and cross-
sectional areas were calculated using the SigmaScan
program (Jandel Scientific Software, St. Rafael, CA,
USA). The other section was embedded in acylacetate
(Entellan, Merck, Darmstadt, Germany) on a glass
slide and used for determination of mineral apposi-
tion rates and bone formation rates by fluorescence
microscopy.15,27 A translucent star-shaped grid was
placed on top of the section with the centre of the star
in the mid-point of the marrow cavity and 16 lines
radiating from the centre point. Since only the cen-
tre of the star was fixed, the 16 radiating lines inter-
sected the periosteal and endocortical surface at ran-
dom. At the point of intersection, the distance between
the middle of the tetracycline label and the middle
of the calcein label was measured at a magnification
200x. An aspect with no labelling was classified as a
point of no mineral apposition. No distance between
the tetracycline and calcein labels was classified as a
point of no measureable mineral apposition, i.e. zero
mineral apposition. The mineral apposition rate was
calculated by dividing the distance between the two
labels by the interlabelling period in days. The lower
limit for detection of a distance between the tetracy-
cline and calcein labels was 5μm, corresponding to a
mineral apposition rate of 0.25 μm per day. The bone
formation rate was the mean mineral apposition rate ×
circumference.
Total serum IGF-I
This was determined by a RIA-assay.28
Statistical analyses
For analysis of differences between the groups the
Kruskal–Wallis test was applied; in case of difference,
the Mann–Whitney U-test was used. P< 0.05 (two-
tailed) was considered statistically significant.29 In the
tables, lower-case letters indicate significance levels at
P<0.05, but >0.01; upper-case letters significance
levels at P<0.01. The following comparisons were
made: 1, the saline-injected group was compared with
the GC-injected group; 2, GHS-injected rats were com-
pared with saline-injected rats; 3, GC-injected rats
were compared with the combined GC +GHS treated
rats; 4, the start control group was compared with
the saline group to delineate changes in the param-
eters in relation to the base line, i.e. start control
group.
RESULTS
GC vs saline
The GC-injected rats had a 4% decrease in BW during
the experimental period (Table 1), and their final
body weight was 14% lower than that of the saline
group. GC had no significant effect on the tibia length
during the experiment (Table 1). A 54% increase in
IGF-I was found in the GC-injected animals compared
with the saline group (Table 1). The DDW of the calf
musculature was reduced by 19% in the GC-injected
rats compared with the saline injected (Table 2), and
the MTT of the calf musculature was reduced by 28%
in the GC-injected rats [Fig. 1(A)]. When the MTT was
normalized to the DDW of the calf musculature a de-
crease was still observed in GC-treated compared with
saline-injected rats [Fig. 1(B)]. No change was found
in the time course of the tetanic tension during 7.5 or
15 seconds of stimulation (Table 2). The GC-injections
resulted in a near total arrest of bone formation rate
[Fig. 2(A)], and a decrease in the periosteal mineralizing
surface [Fig. 2(B)]. The breaking strength was found to
be at the level of the start control group, but did not
differ significantly from the breaking strength of the
saline group (Table 3).
GHS counteracts GC-induced effects on bone 269
Table 1 BW at start, final BW, body weight change, tibia length and total serum IGF-I in 8-month-old female rats injected with growth
hormone secretagogue (GHS) and/or glucocorticoids (GC) for 3 months
Start BW Final BW Change in BW Tibia length Serum IGF-I
Groups n(g) (g) (g) (mm) (ng/ml)
Start control 14 264 ±4−−38.2 ±0.2 497 ±19
Saline 14 264 ±4 294 ±4A30 ±3 38.5 ±0.2 386 ±21A
GC 14 264 ±4 254 ±8a,B −10 ±4B38.2 ±0.2 596 ±51B
GHS 14 264 ±4 313 ±6A,b 49 ±3B38.7 ±0.3 475 ±28b
GC +GHS 14 263 ±4 270 ±6D9±3c,D 38.2 ±0.2 568 ±18D
a: P< 0.05, A: P< 0.01 against the start control group, b: P< 0.05, B: P< 0.01, against the saline injected group, c: P< 0.05 against
the GC-injected group, D: P< 0.01 against the GHS-injected group.
Effect of GHS in normal animals
Injections with GHS increased the body weight by 19%
and the final BW 7% greater than the saline group. The
IGF-I concentrations were increased by 21% compared
with the saline group. GHS had no effect on muscle
mass (DDW) or in tibia length. Likewise, the maximal
tetanic tension [Fig. 1(A)] and the time course of tetanic
tension (Table 2) were unchanged compared with the
saline group. The bone formation rate at the femur
mid-diaphyseal periosteum [Fig. 2(A)] was three times
that of the saline group and there was a small non-
statistically significant increase in mineralizing surface
[Fig. 2(B)]. No significant changes were found in fe-
mur mid-diaphyseal cross-sectional area or breaking
strength (Table 3).
Effect of GHS in GC-injected animals
The group injected with GC and GHS in combina-
tion had a small BW gain in contrast to the group in-
Table 2 Dry defatted weight of the calf musculature and time
course of the tetanic tension during the first 15 seconds of
stimulation in 8-month-old female rats injected with growth
hormone secretagogues (GHS) and/or glucocorticoid (GC) for
3 months. Tetanic tension is given as a percentage of maximal
tetanic tension (MTT)
Tetanic tension Tetanic tension
after stimulation after stimulation
Dry defatted for 7.5 s for 15 s
Groups: nweight (mg) (% of MTT) (% of MTT)
Start Control 14 364 ±10 31.0 ±3.8 12.5 ±1.5
Saline 14 363 ±8 38.7 ±4.6 13.5 ±1.2
GC 14 295 ±8A, B 36.2 ±2.7 16.6 ±1.6
GHS 14 373 ±8 38.5±3.8 15.7 ±1.6
GC +GHS 14 318 ±9D34.8 ±2.8 14.6 ±0.9
A: P< 0.01 against the start control group, B: P< 0.01 against the
saline-injected group, D: P< 0.01 against the GHS-injected group.
jected with GC alone, which had a significant BW loss
(Table 1). The tibia length remained at the same level as
in the start control group. The DDW of the calf muscu-
lature was lower than that of the GHS-infected group
(Table 2). The MMT of animals injected with GC +GHS
in combination was decreased compared with the
saline and GHS treated groups, but increased signifi-
cantly compared with the GC-treated group [Fig. 1(A)].
When the MTT was corrected for differences in mg
DDW of the calf musculature, no difference was found
compared with the saline group [Fig. 1(B)]. No change
was observed in the time course of tetanic tension
(Table 2). The mid-diaphyseal periosteal bone forma-
tion rate was lower compared with the start control
group and the group injected with GHS, but four-fold
higher than the group injected with GC alone [Fig. 2(A).
Likewise, the periosteal mineralizing surface of animals
injected with GC +GHS was clearly increased com-
pared with the rats injected with GC alone [Fig. 2(b)].
This did not, however, result in a significant increase in
the breaking strength of the femur (Table 3). The IGF-I
concentration of the GC +GHS was not significantly
different from that of the GC group (Table 1).
DISCUSSION
Growth hormone secretion decreases with age; after
the age of 60, many elderly subjects have low levels
of GH and IGF-I.30,31 Muscle strength, muscle mass,
bone strength and mass also decrease with aging, and
these losses are further exacerbated by glucocorticoid
treatment. Rudman et al.32 hypothesized that replac-
ing GH in elderly subjects with low levels of serum
GH and serum IGF-I might result in improvements of
physical function; they treated elderly men who had
low serum IGF-I values with GH and found an in-
crease in serum IGF-I, lean body mass and a reduc-
tion in body fat mass.32 Several studies have shown
270 N.B. Andersen et al.
Fig. 1 Maximum tetanic tension (MMT) (A) and MTT normalized to
dry defatted weight (DDW) (B) of the calf muscles in 8-month-old
female rats injected with growth hormone secretagogue (GHS)
and/or glucocorticoid (GC) for 3 months. Mean values with SEM. N:
newton. A: P< 0.01 against the start control group; B: P< 0.01
against the saline group; c: P< 0.05 against the GC-injected
group; D: P< 0.01 against the GHS-injected group.
similar changes in body composition, but it has been
difficult to demonstrate improvements in skeletal mus-
cle strength as a result of the GH treatment.33, 34 Welle
et al.35 found, however, an increase in skeletal muscle
strength after GH treatment for 3 months. GHSs might
have an advantage compared with GH, because some
GHS-preparations can be given orally and some have
been shown to stimulate pulsatile GH secretion.36
In the present study, the GHS-preparation , Ipamore-
lin, injected subcutaneously in rats increased the to-
tal serum concentration of IGF-I by 23% and BW by
7% compared with the saline-injected group, without
significant effect on tibial length. Ipamorelin did not
change skeletal muscle mass or MTT compared with
the saline group. The bone formation rate was clearly
increased in the Ipamorelin-injected rats. The amount
of new bone which corresponded to a 0.1 mm increase
in the thickness of the diaphyseal bone was not suf-
ficient to significantly change the breaking strength
of the bone. In the present study, however, a signifi-
cant stimulation of longitudinal growth was not found.
In the dose–response study of Johansen et al.,25 longi-
tudinal bone growth of 4-month-old rats was signifi-
cantly stimulated by Ipamorelin. In the present study,
the rats were 8–11 months old and had passed the
period of longitudinal growth, even though the epi-
physeal growth plate is still cartilaginous in rats at
that age.
Total serum-IGF-I increased with GC treatment, sup-
porting previous observations.11,37 However, Skjærbæk
et al.38 found a reduced amount of free IGF-I, al-
though the total amount of IGF-I was unchanged or
Fig. 2 Periosteal bone formation rate (A) and mineralizing surface
(B) at the femur mid-diaphysis in 8-month-old female rats injected
with growth hormone secretagogue (GHS) and/or glucocorticoid
(GC) for three months. Mean values with SEM. a: P< 0.05, A:
P< 0.01 against the start control group; B: P< 0.01 against the
saline group; C: P< 0.01 against the GC-injected group; D:
P< 0.01 against the GHS-injected group.
GHS counteracts GC-induced effects on bone 271
Table 3 Mid-diaphyseal cross-sectional area, medullary area and
breaking strength of the femur of 8-month-old female rats injected
with growth hormone secretagogue (GHS) and/or glucocorticoid
(GC) for 3 months
Cortical bone
cross-sectional Medullary Breaking
Groups narea (mm2) area (mm2) strength (N)
Start control 14 6.2 ±0.1 2.7 ±0.1 165 ±6
Saline 14 6.2 ±0.1 2.9 ±0.1 182 ±6
GC 14 6.1 ±0.1 2.5 ±0.1 166 ±6
GHS 14 6.3 ±0.1 2.5 ±0.1 180 ±5
GC +GHS 14 6.1 ±0.1 2.6 ±0.1 175 ±4
increased during GC-treatment. In rats, about 5% of the
IGF-I is free.39 The change in weight induced by GC-
treatment was significantly correlated with free IGF-I
and not with total IGF-I.38 GC-treatment influences
the relationship between IGF-I and its binding pro-
teins. In the present experiment, even though the total
serum IGF-I was increased, the free serum IGF-I may
well be reduced, resulting in reduced IGF-I biological
activity.
The present study supports a possible therapeu-
tic role for GHSs in patients receiving catabolic glu-
cocorticoids and is in agreement with Gertz et al.21
who demonstrated that a GH secretagogue given i.v.
to young men partially reversed glucocorticoid sup-
pression of GH secretion. Likewise, it supports the
study of Lieber et al.19 who immobilized a hind
limb of dogs; GHS given during the remobilization
phase increased muscle strength markedly. Concern-
ing bone, oral administration of GHSs has been shown
to increase markers of bone turn-over in young obese
males40 and in elderly adults.41 A partial attenua-
tion of the GH response to GHS has been shown
i.e. desensitization.25,40,42 The mechanism behind this
phenomenon is not yet clear, but could be due to
negative feedback by GH or IGF-I or to down regu-
lation of the GHS receptor. Despite the decrease in
serum GH, serum IGF-I was increased during these
studies and the desensitization seemed to be partial and
reversible.40 Whether coadministration of GC changes
the desensitization mechanism is unclear. It might be
speculated that the production of immunoglobulins
against Ipamorelin could be lowered with coadminis-
tration of GC possibly affecting its half-life. The normal
half-life of Ipamorelin is 30–60 min in rats after i.v.
administration.43
In conclusion, the decrease in bone formation found
in GC-injected rats can be counteracted by administra-
tion of GHS.
ACKNOWLEDGEMENTS
Novo Nordisk A/S, The Danish Health Research Coun-
cil, grants no. 9500922 (Centre for Molecular Gerontol-
ogy), 9503023 and 9600822 (Aarhus University-Novo
Nordisk Centre for Research in Growth and Regener-
ation), Aarhus University Research Foundation, Novo
Nordisk Foundation. The skilled technical assistance
of C. Knæhus, J. Utoft, E. K. Mikkelsen and linguistic
revision of M. Fischer are gratefully acknowledged.
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... Malmlöf et al., using similar steroid doses (5 mg  kg À1  d À1 ) and Ipamorelin (0.4 or 1.6 mg  kg À1  d À1 ), observed that Ipamorelin normalized GH secretion and IGF-I levels in steroid treated rats [29].The observation that Ipamorelin did not increase IGF-I may indicate GH resistance in the liver as also observed in diabetic mice [28]. In a recent study Ipamorelin was found to counteract the decrease in bone formation by glucocorticoid, and in that study there was no difference in serum IGF-I between the groups studied, either [30]. We did not measure GH levels, though it might have enlightened this observation, as GH may stimulate local IGF-I production which may in turn exert autocrine and paracrine effects. ...
Niels Kristian Aagaard
Thorbjørn Grøfte Jacob Greisen
Hendrik Vilstrup
Growth hormone (GH) reduces the catabolic side effects of steroid treatment via effects on the amino-nitrogen metabolism. Ipamorelin is a synthetic peptide with GH releasing properties. We wished to study the metabolic effects of Ipamorelin and GH on selected hepatic measures of alpha-amino-nitrogen conversion during steroid-induced catabolism. Five groups of rats were included: (1) free-fed controls (2) pair-fed controls (3) prednisolone (delcortol, 4 mg x kg(-1) x day(-1)) (4) prednisolone and GH (1 mg x kg(-1) x day(-1)) (5) prednisolone and Ipamorelin (0.5 mg x kg(-1) x day(-1)). After seven days the hepatic capacity of urea-N synthesis (CUNS) was determined in parallel with measurements of liver mRNA levels of urea cycle enzymes, whole-body N-balance, and N-contents of various organs. Compared to pair-fed controls, prednisolone increased CUNS (p<0.01) as well as the expression of urea cycle genes (p<0.01), and decreased N-balance (p<0.01) as well as organ N-contents (p<0.05). Compared to prednisolone treated animals, co-administration of GH reduced CUNS by 33% (p<0.01), normalized urea cycle gene expression, improved N-balance 2.5-fold, and normalized or improved organ N-contents. In prednisolone treated rats Ipamorelin reduced CUNS by 20% (p<0.05), decreased the expression of urea cycle enzymes, neutralised N-balance, and normalized or improved organ N-contents. Accelerated nitrogen wasting in the liver and other organs caused by prednisolone treatment was counteracted by treatment with either GH or its secretagogue Ipamorelin, though at the doses given less efficiently by the latter. This functional study of animals confirms that the GH secretagogue exerts GH related metabolic effects and may be useful in the treatment of steroid-induced catabolism.
Cory Mayfield
Ioanna K Bolia
Cailan Feingold
Alexander Weber
Background Therapeutic peptides are short-chain amino acids that regulate cellular functions and facilitate biochemical processes. In recent years, there has been significant growth in the global market for therapeutic peptides and thus its popularity among patients. Given the increase in the development of peptides and increased marketing to patients for orthopaedic injuries, it is critical for orthopaedic surgeons to understand the current evidence behind these therapeutic peptides. Purpose To evaluate the current evidence and applications of injectable peptide therapy, focusing on its potential in regenerative medicine and sports performance, to help orthopaedic providers better understand the current state of different therapeutic peptide approaches. Study Design Narrative review. Methods A comprehensive literature search was conducted using PubMed to identify biochemical and clinical studies on the most popular types of injectable peptide therapy. Key peptides evaluated included BPC-157, TB-4, TB-500, CJC-1295 + ipamorelin, tesamorelin, and GHK-Cu. Results BPC-157 demonstrated potential benefits in tendon and muscle repair, but these findings are largely unvalidated in human trials. A single human case series reported improvements in pain after intra-articular knee injections of BPC-157, although significant methodological flaws and a lack of controls limit its applicability and reliability. TB-4 and its derivative TB-500 promoted angiogenesis and tissue repair in preclinical models, but human orthopaedic data are lacking, and both remain banned substances in sports. CJC-1295 combined with ipamorelin showed significantly improved maximum tetanic tension in murine models with glucocorticoid-induced muscle loss, but these findings are limited to animal studies. Tesamorelin, approved for treating HIV-associated lipodystrophy, has no supporting orthopaedic evidence. GHK-Cu showed promise in wound healing and anti-inflammatory effects, but no clinical data support its use for musculoskeletal conditions. Conclusion While peptide therapy may possess significant therapeutic and regenerative potential, it is critical that orthopaedic and sports medicine providers understand the current lack of evidence to support the clinical use of these peptides. Importantly, information regarding the indications, dosing, frequency, and duration of treatment remains unknown. Despite the popularity of these peptides in mainstream media and among patients, significant research regarding the safety and efficacy of these therapeutic methods is required before definitive recommendations can be made to patients.
Philip A Carpino
The 28 amino acid polypeptide, ghrelin, is a novel neuroendocrine hormone synthesised primarily in the stomach. It stimulates growth hormone (GH) secretion, increases food intake and appetite and promotes body weight gain by decreasing fat mass metabolism, particularly in rodent models. The pharmacological properties of this peptide are mediated by the type 1a growth hormone secretagogue receptor (GHS-R1a), a G-protein-coupled receptor localised predominantly in the pituitary gland and hypothalamus. The discovery of new agents that either mimic or modulate the actions of ghrelin via the GHS-R1a has attracted considerable interest in recent years. Many peptidyl and peptidomimetic GHS-R1a agonists such as growth hormone-releasing peptides (GHRPs) and growth hormone secretagogues (GHSs) that stimulate GH secretion and increase plasma levels of insulin-like growth factor-1 (IGF-1) have been identified. First generation GHSs have undergone extensive clinical evaluation in humans, but have shown disappointing results in the treatment of diseases such as frailty, the age-related decline in body composition. Several of these compounds were found to increase adiposity in rodent models. Second generation dipeptidyl and third generation small molecule GHSs have recently been profiled in various animal models. A third generation of GHS with partial agonist activity at the GHS-R1a showed anabolic activity in rodent models, increasing fat-free (muscle) mass rather than fat mass. A few GHS-R1a receptor antagonists have been disclosed, although these compounds have exhibited poor selectivities or weak affinities for the receptor. Since endogenous ghrelin appears to play an important role in the long-term regulation of energy balance, GHS-R1a antagonists may be useful in the prevention of weight gain, following weight loss, or in the treatment of obesity, a pathological condition characterised by excess adiposity. This review describes the pharmacology of select ghrelin receptor modulators and highlights new ghrelin receptor agonists and antagonists that have appeared in the literature from 1999 – 2002.
Ernest A. Adeghate
Abdul S Ponery
To examine the effect of ipamorelin (IPA), a novel pentapeptide with a strong growth hormone releasing potency, on insulin secretion from pancreatic tissue fragments of normal and diabetic rats. Diabetes mellitus was induced by streptozotocin (60 mg kg(-1)). Four weeks after the induction of diabetes, pancreatic tissue fragments of normal and diabetic rats were removed and incubated with different concentrations (10(-12) - 10(-6) M) of IPA. Insulin release from the pancreas was measured by radioimmunoassay. Ipamorelin evoked significant (p<0.04) increases in insulin secretion from the pancreas of normal and diabetic rats. Either diltiazem or yohimbine or propranolol or a combination of atropine, propranolol and yohimbine inhibited IPA-evoked insulin secretion significantly (p<0.03) from the pancreas of normal and diabetic rats. Atropine caused a significant (p<0.007) reduction in the IPA-induced insulin secretion in diabetic but not in normal rats. IPA stimulates insulin release through the calcium channel and the adrenergic receptor pathways. This is the first study to examine the effect of ipamorelin on insulin secretion in the pancreas.
A Juul Peter Bang
Niels Thomas Hertel N E Skakkebaek
Serum levels of insulin-like growth factor-I (IGF-I) increase with age and pubertal development. The large variation in circulating IGF-I levels in adolescence makes it difficult to use the IGF-I value of a single child in the assessment of his growth status. In addition, the interference of IGF-binding proteins in many IGF-I assays contributes to this problem. We measured IGF-I in acid-ethanol-extracted serum from 1030 healthy children, adolescents, and adults, employing a RIA that reduces interference of IGF-binding proteins by using monoiodinated Tyr31-[125I]des-(1-3)IGF-I as radioligand. Mean serum IGF-I concentrations increased slowly in prepubertal children from 80-200 μgs/L with a further steep increase during puberty to approximately 500 μgs/L. After puberty, a subsequent continuous fall in circulating IGF-I levels was apparent throughout adulthood to a mean of 100 μgs/L at the age of 80 yr (P < 0.0001). Girls had maximal IGF-I levels at 14.5 yr of age, whereas boys had peak IGF-I levels 1 yr later. This is almost 2 yr later than average peak height velocity. The large variation in serum IGF-I levels during puberty was diminished when data were separated according to sex and Tanner stage of puberty. Interestingly, we found a significant variation with age within the Tanner stages; there was an increase in serum IGF-I concentrations with age in the early pubertal stages and a decrease in the late stages (P < 0.05). Serum IGF-I increased concomitantly with increasing testicular volume. Multiple regression analysis revealed that serum IGF-I levels predicted height velocity in the following year (r = 0.33; P < 0.0001). Body mass index did not correlate significantly with serum IGF-I in prepubertal children in a multiple regression analysis. In conclusion, there was a significant variation in serum IGF-I levels with age within a given Tanner stage of puberty in addition to the well known increase with increasing age or pubertal stage. Accordingly, the effects of sex, age, and puberty on serum IGF-I cannot be separated into simple additive components when studying 1030 children in a cross-sectional design. Thus, the age-, sex-, and puberty-corrected IGF-I values may, in fact, improve the use of serum IGF-I as a diagnostic tool to distinguish between a child with retarded puberty and a GH-deficient individual.
Ian Chapman
Mark Bach
Eve Van Cauter
Michael O Thorner
Aging is associated with declining activity of the GH axis, possibly contributing to adverse body composition changes and increased incidence of cardiovascular disease. The stimulatory effects on the GH-insulin-like growth factor I (IGF-I) axis of orally administered MK-677, a GH-releasing peptide mimetic, were investigated. Thirty-two healthy subjects (15 women and 17 men, aged 64-81 yr) were enrolled in a randomized, double blind, placebo-controlled trial. They received placebo or 2, 10, or 25 mg MK-677, orally, once daily for 2 separate study periods of 14 and 28 days. At baseline and on day 14 of each study period, blood was collected every 20 min for 24 h to measure GH, PRL, and cortisol. Attributes of pulsatile GH release were assessed by 3 independent algorithms. MK-677 administration for 2 weeks increased GH concentrations in a dose-dependent manner, with 25 mg/day increasing mean 24-h GH concentration 97 +/- 23% (mean +/- SE; P < 0.05 vs. baseline). This increase was due to an enhancement of pree...
Stephen Welle
Colleen Thornton M Statt B McHenry
The rate of synthesis of myofibrillar proteins is slower in muscle of healthy subjects over 60 yr old than it is in young adults. Previous research suggests that reduced activity of the GH/insulin-like growth factor-I system could be a determinant of this slowing of protein synthesis. To test the hypothesis that GH could rejuvenate the rate of myofibrillar protein synthesis, we studied healthy subjects over 60 yr old, after a single injection (0.03 mg/kg.sc) of recombinant human GH (n = 6 males/2 females) or placebo (n = 6 males/2 females), or after 3 months of either GH (0.03 mg/kg, sc, 3 x /week, n = 5 males) or placebo (n = 5 males) treatment. Myofibrillar protein synthesis and whole-body protein metabolism were evaluated with the tracer L-[1-13C]leucine. GH reduced whole-body leucine oxidation by 36% (P < 0.01) in the single injection study. There was no effect of GH on whole-body protein breakdown or synthesis, or on myofibrillar protein synthesis in the quadriceps, either acutely or after 3 months of treatment. GH treatment for 3 months increased lean body mass (3.3 +/- 0.7 kg, P < 0.01, as evaluated by 40K counting), muscle mass (3.3 +/- 1.1 kg, P < 0.02, as evaluated by urinary creatinine excretion), and thigh strength (14 +/- 5%, P < 0.05, as evaluated by isokinetic dynamometry). We conclude that GH can increase muscle mass and strength in healthy men over 60 yr old, but does not restore a youthful rate of myofibrillar protein synthesis.
H. Oxlund
The effects of a combination of mild exercise and GH injections on bone were studied in old female rats. Biosynthetic human GH, 2.7 mg/kg/day, was injected sc for 73 days. Exercised rats ran 8 m/min on a treadmill for 1 h/day. All rats (age 21 months old) were labeled with a tetracycline injection 56 days and a calcein injection 11 days before killing. The GH injections resulted in an 11-fold increase in femoral middiaphyseal bone formation rate and a 12% increase in cross-sectional area compared with the saline-injected group. The mild exercise doubled the mineralizing surface but did not influence the bone formation rate significantly. The combination of GH injections plus exercise, however, resulted in a further increase of 39% in bone formation rate, primarily at the anterolateral aspects, and an increase of 5% in cross-sectional area compared with the group injected with GH only. The femur ultimate breaking load was increased by 37% and the stiffness by 42% in the group injected with GH compared with the saline-injected group. Exercise alone did not influence the femur mechanical properties. The combination of GH injections plus exercise induced a 4% further increase in ultimate breaking load and 7% further increase in stiffness compared with the group injected with GH alone. The GH injections induced a 117% increase in serum insulin-like growth factor I. The GH-insulin-like growth factor I axis stimulates recruitment of osteoblast precursor cells, resulting in increased bone formation at the periosteal surface. GH injections and mild excercise in combination modulate and increase further the formation and strength of cortical bone in old female rats.
J Svensson
Growth hormone (GH) is of importance for normal bone remodelling. A recent clinical study demonstrated that MK-677, a member of a class of GH secretagogues (GHSs), increases serum concentrations of biochemical markers of bone formation and bone resorption. The aim of the present study was to investigate whether the GHSs, ipamorelin (IPA) and GH-releasing peptide-6 (GHRP-6), increase bone mineral content (BMC) in young adult female rats. Thirteen-week-old female Sprague-Dawley rats were given IPA (0.5 mg/kg per day; n=7), GHRP-6 (0.5 mg/kg per day; n=8), GH (3.5 mg/kg per day; n=7), or vehicle administered continuously s.c. via osmotic minipumps for 12 weeks. The animals were followed in vivo by dual X-ray absorptiometry (DXA) measurements every 4th week. After the animals were killed, femurs were analysed in vitro by mid-diaphyseal peripheral quantitative computed tomography (pQCT) scans. After this, excised femurs and vertebrae L6 were analysed by the use of Archimedes' principle and by determinations of ash weights. All treatments increased body weight and total tibial and vertebral BMC measured by DXA in vivo compared with vehicle-treated controls. However, total BMC corrected for the increase in body weight (total BMC:body weight ratio) was unaffected. Tibial area bone mineral density (BMD, BMC/area) was increased, but total and vertebral area BMDs were unchanged. The pQCT measurements in vitro revealed that the increase in the cortical BMC was due to an increased cross-sectional bone area, whereas the cortical volumetric BMD was unchanged. Femur and vertebra L6 volumes were increased but no effect was seen on the volumetric BMDs as measured by Archimedes' principle. Ash weight was increased by all treatments, but the mineral concentration was unchanged. We conclude that treatment of adult female rats with the GHSs ipamorelin and GHRP-6 increases BMC as measured by DXA in vivo. The results of in vitro measurements using pQCT and Archimedes' principle, in addition to ash weight determinations, show that the increases in cortical and total BMC were due to an increased growth of the bones with increased bone dimensions, whereas the volumetric BMD was unchanged.
Leland Graves
Barbara Lukert
Glucocorticoid-induced osteoporosis is the leading cause of medication-induced osteoporosis. The fracture incidence after 1 yr of glucocorticoid therapy has been found to be 17% and observational studies suggest that fractures eventually occurin 30 to 50% of chronic glucocorticoid-treated patients. An increased risk of fracture develops within as quickly as 3 mo, and with doses of prednisone as low as 2.5 mg daily. Despite effective methods of prevention and treatment, only 50% of chronic glucocoriticoid-treated patients undergo any evaluation for osteoporosis and less than one in four receives treatment. This article addresses the various issues the clinician faces in managing patients receiving chronic glucocorticoids. It begins and ends with a case perspective. Evidence-based information is used to outline the current best strategy for the prevention and treatment of glucocorticoid-induced osteoporosis.