Page 2 - Gastric pentadecapeptide BPC 157

aids in healing of different wounds (i.e.. skin [17], cor- Eiwlircilioii cJ/' rIi:/ic.ii crritl rcwir.i'r:l. cJ/' Ac./ii//c,s rivicloii jiillowiiig irciiisi'c-
neal [S], colon-colon anastomosis [3 I], deep skin burn /kill
[ 121, segmental osteoperiosteal bone defects [ 161). Also, Fiiiiclioiicil iw~/iici/iii~i. Ac/ii//i.s ,/ioiciioiio/ iiic1c.s
it has an angiogenic effect [21] and modulates NO-syn- In general. the iissessment is carried out according the procedure
thesis [19]. This may directly affect repair of connective previously described in details [I]. Briefly. the rats with hind paws
tissues [6], as an ordered multistage process involving moistened. iirc allowed to walk down a confined runway. The foot-
prints were scanned. and measured by computer imaging using a
inflammation and oxidative stress. Since oxidative stress special program from VAMSTECH. Zagreb, Croatia. Measurements
with consequent lipid peroxidation and production of (the distnnce to the opposite foot (TOF). print length (PL). the dis-
HNE has an important role in pathophysiology of tis- tance between the second and fourth or intermediary toes (IT)) are
made from each walking track defining particular parameters (de
sue damage, inflammation and cell growth regulation Medinaceli's print length factor (PLF): toe-spread factor (TSF): in-
[29,30], we investigate the influence of BPC 157 on the termediary toe-spread factor (ITF)) included in AFI-calculation
growth of cultured .tendocytes obtained from the rat (AFI =74(PLF)+ 161(TSF)+48(ITF)-5). follows: PLF=(NPL-
EPL)/EPL: TSF = (ETS-NTS)/NTS; ITF = (EIT-NlT)/NlT. where N
Achilles tendon, quiescent or treated in vitro with HNE. is the norniiil uninjured side. E is the experimental. injured side, and
A comparison to transforming growth factor beta TS is total spreading).
(TGF-P) a factor known to increase matrix expression Tiw.viourc,/,:l. iiii,i,.s/i,~ci/ioii
[ 141 was done. Measurement of failure load (N). failure load per area (Nlmni')
Besides, this gastric pentadecapeptide BPC I57 likely and Young's modulus of elasticity (E) (as an index of elastic length
controls functions of collagen fragments [ 12,231. Like- deformity (Nlnini'). that characterizes tendon resistance to elastic
wise, it has special relation over bone morphogenetic deformation during distension. and also means increased stiffness as
well iis strength. according to formula E = (F/S)/A//lo). where F-
proteins [ 12,231. High stability [8. 12,16,I7.19%2 1,281 force (N). S cross-sectionol area (nim?). A/ elongation of tendon
(i.e., no degradation in human gastric juice even for 24 length (nim). lo tendon length (mm)) is carried out in separate cx-
periments. immedintely after sacrifice. at day 7. 10 and 14. in generally
h, unlike rapidly degraded h-TGF, and h-EGF) [26]) iis described beforc in our recent report [I?] and along with Best and
permits no carrier used in previous [8,I2,16,17.19- coworkers [2] by ii special device Statimat M (Textechno. Germany,
21,281 and present studies. Together, this means that DIN 51221. DIN 53834. IS0 2062 and all equipment of Faculty of
unlike limited delivery of other peptides [ 1.7,9.23] this Textile Technology University of Zagreb. Croatia) using procedure for
tendon bioniechanical assessment described in details by Nikolic and
stable pentadecapeptide is highly resistant to otherwise Hudec [ 151. with special mechanical device (Faculty of Textile Tech-
inescapable degradation of peptides. Likewise, acting nology University 01' Zagreb. Croatia) for sample fixation. Besides. to
alone, withollt carrier, it presents a healing potential of avoid period elapsed between sacrifice and assessment. and the pro-
cedure related to minimizing tissue destruction [2]. we performed as-
its own [8,12,16,17,19-21,311, and may be suitable for sessment immediately after sacrifce to minimize the outconie from
therapy of complex structures such as the Achilles physiologic volues. The data were analyzed automatically by a con-
tendon. nected computer system.
I
,voc~,o.v~'fl/~~' ~l.s.sc'.s.~111171
Inimediately after sacrilice. to avoid problems.related to knee and
Materials and methods ankle flexion angles. the specimen (tendon calcaneal insertion, distal
and proximal tendon part. together with I cin of gastrocnemius and
Drugs soleus muscle complex) is carefully removed. placed at the flat board
and the size of the defect (longest diameter. mm) between the cut ends
Pentadecapeptide BPC 157 (GEPPPGKPADDAGLV. M.W. 1419) of Achilles calculated in a blinded fashion.
(manufactured by Diagen. d.0.o.. Ljubljana. Slovenia) is ii piirtial of
sequence of human gastric juice protein BPC. freely soluble in water at Mic~,.osc.ll?,. ci.v.vl'.vsll~l'lll
pH 7.0 and in saline. It was prepared iis described before The tendons are lixed in buRered formalin (pH 7.4) for 24 h. de-
[8.12,16.17,19-21.31]. Peptide with 99% (HPLC) purity ( I-des-Gly hydrated. and embedded in paraltin wax. Coronal sections of the
peptide as impurity). dissolved in saline was used in all of the experi- midsubs!ance of the tendon :ire cut. stained with hematoxylin and
ments [8,12,16.17,19-21.31]. TGF-Dl was obtained from Sigma. USA. eosin. or Gomori silver stain (reticdin) and Van Gieson andlor Mal-
HNE was kindly given by Institute of Biochemistry Medical Faculty lory trichrome staining (collagen). and examined in a blinded lbshion
University of Graz. Graz. Austria. [2]. At least three step sections per tendon were done. To identify an
early effrct. iit ii critical one day period. we assessed the number of
Esperiiiienrcil procecliire granulocytes and mononuclears. At latter time intervals other pa-
rameters were analyzed microscopically. Reticulin (day 4) as well as
Male Wistar Albino rats. 280-320 g body weight. randomly iis- collagen proportion (days 10 and 14) per visual field were assessed
signed (20 rats per each of groups. 10 for histology only. per each of accordingly with our recent report [ 121 and general methodological
the tested intervals) are used in all of the experiments. All of the ex- consideration. The analyzed area included three high power field.
periments are approved by local Ethic Committee. Under light ether objective 40. next to the tendon resection margin taking into iiccount
anaesthesia. using a previously described procedure [2]. the right regularly more advanced hceling process. and its clear identification in
Achilles tendon is transected transversely 0.5 cin proximal to its all animals [IZ]. Fibroblasts were counted on six power field in cor-
calcaneal insertion. A large tendon defect appears between cut ends. responding area on HE slides [IZ]. For morphonietrical analysis special
Sham procedure included skin lesion only. and non-operated healthy program SFORM. VAMSTECH-Software Company (VAMSTECH.
rats are used as an additional control (since not different these data Zagreb. Croatia) was used.
were not specifically shown). Agents (Ikg b.w.. i.p.. once time daily)
(BPC 157 (dissolved in saline, with no carrier addition) ( 10 pg. 10 ng or Ci4l ld/iirl,.\
10 pg) or saline (5.0 ml)), were firstly applied at 30 niin after surgery.
the last application at 24 h before autopsy. In generally. the experi- Achilles tendons of five sacriliced Wistar rats were taken under
ments were carried out under blind condition. sterile conditions and dissected into smaller pieces (<I mm') treated as