Stable Gastric Pentadecapeptide BPC 157 and Wound Healing

Abstract

Significance: The antiulcer peptide, stable gastric pentadecapeptide BPC 157 (previously employed in ulcerative colitis and multiple sclerosis trials, no reported toxicity (LD1 not achieved)), is reviewed, focusing on the particular skin wound therapy, incisional/excisional wound, deep burns, diabetic ulcers, and alkali burns, which may be generalized to the other tissues healing. Recent Advances: BPC 157 has practical applicability (given alone, with the same dose range, and same equipotent routes of application, regardless the injury tested). Critical Issues: By simultaneously curing cutaneous and other tissue wounds (colocutaneous, gastrocutaneous, esophagocutaneous, duodenocutaneous, vesicovaginal, and rectovaginal) in rats, the potency of BPC 157 is evident. Healing of the wounds is accomplished by resolution of vessel constriction, the primary platelet plug, the fibrin mesh which acts to stabilize the platelet plug, and resolution of the clot. Thereby, BPC 157 is effective in wound healing much like it is effective in counteracting bleeding disorders, produced by amputation, and/or anticoagulants application. Likewise, BPC 157 may prevent and/or attenuate or eliminate, thus, counteract both arterial and venous thrombosis. Then, confronted with obstructed vessels, there is circumvention of the occlusion, which may be the particular action of BPC 157 in ischemia/reperfusion. Future Directions: BPC 157 rapidly increases various genes expression in rat excision skin wound. This would define the healing in the other tissues, that is, gastrointestinal tract, tendon, ligament, muscle, bone, nerve, spinal cord, cornea (maintained transparency), and blood vessels, seen with BPC 157 therapy.

Keywords: bleeding disorder; concept practical applicability; fistula; stable gastric pentadecapeptide BPC 157; wound healing.

FIGURE 1

Burn skin lesions in mice and BPC 157 therapy effect. The effects of the gastric pentadecapeptide BPC 157 were investigated on deep partial skin thickness burns (1.5 × 1.5 cm) covering 20% of the total body area, when administered topically or systemically in burned mice (Mikus et al., 2001). Characteristic wound presentation at one week after injury, grossly, the poor healing in the untreated control or mice treated with vehicle only (c (black letter)) was completely reversed in BPC 157 cream–treated mice (b (black letter)) (1 μg/g neutral cream thin layer once time daily). Likewise, BPC 157 mice exhibited an increased breaking strength and relative elongation of burned skin and reduced water content in burned skin. Contrarily, silver sulfadiazine regimen did not achieve these healing effects. Microscopically (lower), at the postinjury day 3, in control mice, the burned area exhibits severe edema in the dermis and subcutis as well as an exudate with abundant edematous fluid on the surface (white arrow). Coagulated blood vessel walls and a proportion of vessels with fibrin clots (dashed white arrow) (HE, x4 (c), x10 (C)). Contrarily, BPC 157 mice have much less pronounced edema (black arrow), weak cellular infiltrate, and exudate, and the blood vessels walls seem to be more preserved (dashed black arrow) and in more vessels, endothelial cell can be observed. Almost no arterial clots (HE, x4 (b), x10 (B)) were observed.

FIGURE 2

Excisional wound in diabetic rats (0, 1) and in rats with ligation of the right iliac artery and vein (2) (upper) and BPC 157 therapy (lower). At 3 days before wounding, alloxan (300 mg/kg sc), thin layer of the BPC 157 cream (1 µg/1 g neutral cream) (white number), or neutral cream (black number) was immediately given upon wounding (0, 0). Advanced wound healing presentation at 24 h in BPC 157 rats (1, white), but not in controls (1, black). Likewise, advanced healing in the rats with ligated right iliac arteries and veins, when ingested BPC 157 through drinking water (10 μg/kg, 10 ng/kg, 0.16 μg/ml, 0.16 ng/ml, and 12 ml/rat/day) at postsurgery day 4 (2, white), but not in controls (2, black).

FIGURE 3

Tracheocutaneous fistulas and BPC 157 therapy. After injury induction, BPC 157 dissolved in saline (10 μg, 10 ng/kg body weight) given per-orally in drinking water till the sacrifice (0.16 μg/ml, 0.16 ng/ml, and 12 ml/day/rat). At postsurgery day 7, fistula closure, closed tracheal defect, and closed skin defect (B, b) (arrow) were observed in BPC 157 rats. Contrarily, in controls, fistula remained open, fistulous channel was formed in the skin, and open tracheal defects were observed (C, c) (arrow) (HE, x4).

FIGURE 4

BPC 157 and fistulas closing. External (A–D) and internal (E, F) fistulas. External fistulas. (A) Persistent esophagocutaneous fistula and BPC 157 therapy effect. BPC 157 was given per-orally, in drinking water (10 μg/kg, 10 ng/kg, i.e., 0.16 μg/ml, 0.16 ng/ml, 12 ml/rat/day) until sacrifice, or intraperitoneally (10 μg/kg, 10 ng/kg) with first application at 30 min after surgery, last at 24 h before sacrifice. To establish NO-system involvement, L-NAME (5 mg/kg i.p.) (worsening) and/or L-arginine (100 mg/kg i.p.) (beneficial effect) were given alone or together; first application at 30 min after surgery, last at 24 h before sacrifice. BPC 157 (10 μg/kg, i.p. or p.o.) given with L-NAME (5 mg/kg i.p.) and/or L-arginine (100 mg/kg i.p.) and it maintained its original beneficial effect. A closely interrelated process of unhealed skin, esophageal defects, unhealed fistulas (upregulated eNOS, iNOS, and COX2 mRNA levels), usually lethal, particularly NO-system–related and therapy dependent, illustrate a largely open skin defect in controls at day 3 (c) and a closed skin defect in BPC 157 rats (b) (Ceserec et al., 2013). (B) Initial presentation of the persistent gastrocutaneous fistula and BPC 157 therapy effect. Huge gastrocutaneous fistula leaking at 1st postoperative day in control rats (c) and dried fistula without any leakage in BPC 157 rats (b (Skorjanec et al., 2009). The rats received pentadecapeptide BPC 157 (0.16 μg/ml) in drinking water (12 ml/rat) until sacrifice or drinking water only. A comparative study of the BPC 157 beneficial effect was done with intraperitoneal application, once daily, intraperitoneally (per kg body weight) 10 μg, 10 ng, or 10 pg BPC 157, while standard agents 10 mg atropine, 50 mg ranitidine, and 50 mg omeprazole provide only a weak effect. 6-alpha-methylprednisolone (1 mg/kg intraperitoneally, once daily) was given alone which produced a considerable worsening and was completely eliminated with coadministration of BPC 157 10 μg/kg intraperitoneally. (C) Persistent duodenocutaneous fistula and BPC 157 therapy effect. BPC 157 was given per-orally, in drinking water (10 μg/kg, 10 ng/kg, i.e., 0.16 μg/ml, 0.16 ng/ml, and 12 ml/rat/day) till sacrifice, or alternatively, 10 μg/kg and 10 ng/kg intraperitoneally; first application at 30 min after surgery, last at 24 h before sacrifice. To establish a connection with the NO-system, l-NAME (5 mg/kg intraperitoneally) (worsening) and/or L-arginine (100 mg/kg intraperitoneally) (beneficial effect) were given alone or together; first application at 30 min after surgery, last at 24 h before sacrifice. BPC 157 10 μg/kg, intraperitoneally or per-orally, was given with l-NAME (5 mg/kg intraperitoneally) and/or L-arginine (100 mg/kg intraperitoneally) and it maintained its original beneficial effect. Controls simultaneously received an equivolume of saline (5.0 ml/kg intraperitoneally) or water only. Duodenal fistula leaking through skin defect and still open duodenal defect at 2 weeks following fistula creation by anastomosis between the skin and duodenum defect (c). Closed both skin and duodenal defect in BPC 157 rats (fistula closed, b) (Skorjanec et al., 2015). (D) Persistent colocutaneous fistula and BPC 157 therapy effect. BPC 157 accelerated parenterally or per-orally the healing of colonic and skin defect, leading to the suitable closure of the fistula, macro/microscopically, biomechanically, and functionally (larger water volume sustained without fistula leaking) (Klicek et al., 2013). In anesthetized rats, we created the colocutaneous fistula at 5 cm from the anus, colon defect of 5 mm, and skin defect of 5 mm. The rats received pentadecapeptide BPC 157 (0.16 μg/ml) or nothing in the drinking water (12.0 ml/rat) until the sacrifice or once daily, intraperitoneally BPC 157 10.0 μg/kg, 10.0 ng/kg, or saline (5.0 ml/kg b.w.); first application at 30 min after surgery, final 24 h before sacrifice. For comparison, sulfasalazine (50 mg/kg intraperitoneally, once daily) (moderately effective) or 6-α-methylprednisolone (1.0 mg/kg intraperitoneally, once daily) (aggravation) was given. To establish connection with the NO-system, L-NAME (5.0 mg/kg) (worsening) and L-arginine (200.0 mg/kg) (effective only with blunted NO-synthesis, but not alone) were given intraperitoneally alone or in combination (d-arginine 200.0 mg/kg was not effective, data not shown). BPC 157 given with NO-agents, which maintained its original effect. Initial colon defect presentation at 4 weeks following fistula creation by anastomosis between the skin and colon defect (c, middle). Presentation after next 2 weeks (c, b): in control rats, drinking water was continuously given (12 ml/day/rat) (defecation through fistula, c) and in BPC 157 rats, BPC 157 (10 μg/kg/day) was given in drinking water (0.16 μg/ml/day/rat) (fistula closed, b). Internal fistulas. (E) Persistent colovesical fistula and BPC 157 therapy effect. With internal fistulas in the colon and the bladder, with BPC 157 therapy, the colon and bladder defects showed simultaneous healing effects, including closing of the colovesical fistula in a matching healing process (Grgic et al., 2016). BPC 157 was given per-orally in drinking water (10 μg/kg, 12 ml/rat/day) until sacrifice, or 10 μg/kg or 10 ng/kg was given intraperitoneally once daily, with the first application at 30 min after surgery and the last application at 24 h before sacrifice. The controls simultaneously received an equivolume of saline (5.0 ml/kg ip) or water only (12 ml/rat/day). At postoperative day 28, voiding through fistula in controls (fecaluria) (c) and a normal voiding in BPC 157-rats (b). (F) Persistent rectovaginal fistula and BPC 157 therapy effect. We suggest BPC 157 healing of the rats’ rectovaginal fistulas (since spontaneous only poor healing as those in humans) as a realization of the internal fistula-healing concept, an efficient “wound-healing capability” as the therapy of the complicated internal fistula healing. BPC 157 was given per-orally, in drinking water (10 μg/kg or 10 ng/kg, 0.16 μg/ml, or 0.16 ng/ml 12 ml/rat/day) till sacrifice, or alternatively, 10 μg/kg and 10 ng/kg intraperitoneally once daily; first application at 30 min after surgery, last at 24 h before sacrifice. Controls simultaneously received an equivolume of saline (5.0 ml/kg ip) or water only (12 ml/rat/day). At postoperative day 21, defecation through vagina in controls (c) and a normal defecation in BPC 157 rats (b) (Baric et al., 2016).

FIGURE 5

Hematoma formation following tibial diaphysis fracture and BPC 157 therapy effect, in analogy with accomplished all four events (vessel constriction, the primary platelet plug, the secondary plug, and resolution of the clot) that occur in a set order following the loss of vascular integrity (Stupnisek et al., 2012) involved in the wound healing. We suggest BPC 157 healing starting with a rapid formation of the adequate hematoma as a connective scaffold between the stumps. BPC 157 (10 ng/kg) was given as a 1-ml bath to the injury, immediately after injury induction. Controls simultaneously received an equal of saline as a bath to the injury. At 1 min after application, hematoma within fracture gap (b) further progressed at 2 min (B) in BPC 157 rats. Diffuse bleeding in controls at 1 min (c) and 2 min (C), weak hematoma formed outside of the fracture gap.

FIGURE 6

Perforated cecum defect and BPC 157 therapy effect. i. Presentation of the perforated cecum defect (o-immediately after perforation, before therapy) as an illustration of the rapid healing effect immediately after wounding, with vessels “running” toward the defect augmented by BPC 157 bath application (10 μg/kg) (USB microcamera). b—vessels recruitment presentation immediately under the immersion of the BPC 157 bath, which had been applied at the cecum, and presentation immediately thereafter →b) with corresponding controls (saline bath 1 ml/rat) presentation c, →c). ii. Resultant bleeding from the perforated defect (C (controls), decreased in BPC 157 rats (B). iii. Final failure of the perforated defect healing in controls (C) (postinjury day 7) and completely healed defect in BPC 157 rats (B). This beneficial effect goes along with counteraction of the worsening effect of both NOS-blocker L-NAME (5 mg/kg), or NOS substrate L-arginine (100 mg/kg) (directly applied to the perforated cecum, alone or combined, and spread through the abdominal cavity), and normalization of the increased MDA- and NO-values in the cecum (Drmic et al., 2018).

FIGURE 7

Venous occlusion and BPC 157 therapy. Given BPC 157 (as an abdominal bath) immediately before venography, at a particular point, venography demonstrated a rapid recruitment of the collaterals to bypass occlusion and reestablish blood flow. In the rats with infrarenally occluded inferior caval vein, venography in the inferior caval vein below the ligation shows that the left ovarian vein is rapidly presented as the major pathway. The other veins (such as epigastric veins, intercostal veins, mammary veins, iliolumbar veins, paraumbilical vein, azygos vein, and right ovarian vein) accordingly appear in BPC 157 rats (B), unlike in controls (C). Together, this means rapidly activated efficient compensatory pathways and the ligation-stop at the inferior caval vein efficiently bypassed (Vukojevic et al., 2018). Both kidneys and canal systems and confluence of the inferior caval vein to the right heart demonstrated that redistribution of otherwise trapped blood volume was rapidly achieved (Vukojevic et al., 2018). In the rats with occluded superior mesenteric vein, occlusion was made at the end of the superior mesenteric vein. Venography in superior mesenteric vein below the ligation shows that bypassing through inferior anterior pancreaticoduodenal vein and superior anterior pancreaticoduodenal vein to the pyloric vein toward the portal vein rapidly occurs in BPC 157 rats (b), unlike failed bypassing presentation in controls with occluded superior mesenteric vein (c).

FIGURE 8

Gene expression analysis and BPC 157 therapy. With the therapy done immediately after wounding, we performed the Akt1, Braf, Egfr, Egr1, Grb2, Hdac7, Kras, Mapk1, Mapk3, Mapk14, Nos3, Pik3cd, Plcg1, Prkcg, Ptk2, Pxn, Src, Srf, and Vegfa genes expression analysis (○, no significant change in gene expression; ▲, increased gene expression) in the rats’ excision wound and in the skin and subcutaneous tissue, done at 2, 5, and 10 min following BPC 157 application. Thus, considering the quite rapid presentation of the BPC 157 beneficial effect in the wound healing, it is likely indicative that the expression of all of the genes Akt1, Braf, Egfr, Egr1, Grb2, Hdac7, Kras, Mapk1, Mapk3, Mapk14, Nos3, Pik3cd, Plcg1, Prkcg, Ptk2, Pxn, Src, Srf, and Vegfa is increased at the 10-min interval. An additional indicative point is sequential involvement. The increased expression of the Akt1, Grb2, Nos3, Pik3cd, Prkcg, Ptk2, and Src appears immediately at 2-min interval. Braf, Egfr, Egr1, Hdac7, Mapk1, Mapk3, Mapk14, Plcg1, Prkcg, Ptk2, Pxn, Src, Srf, and Vegfa increased expression is noted at 5 min. The increased expression of the Kras appears at 10 min. Of note, an enormous number of the interactions between the genes involved prevents a more definitive understanding of function insight outcome. However, the evidence is obtained that BPC 157 action initially affects expression of particular genes (i.e., Akt1, Grb2, Nos3, Pik3cd, Prkcg, Ptk2, Src). Then, it involves other set of the genes (Braf, Egfr, Egr1, Hdac7, Mapk1, Mapk3, Mapk14, Plcg1, Prkcg, Ptk2, Pxn, Src, Srf, and Vegfa). Subsequently, it concludes with additional genes set (i.e., Kras). Together, these may likely represent the complex way how the action, which will eventually resolve the lesion, may start and progress.

FIGURE 9

(Left, 14 days) (right, 28 days). Transected muscle and BPC 157 therapy (Staresinic et al., 2006). Quadriceps muscle was completely transected transversely 1.0 cm proximal to the patella means a definitive defect that cannot be compensated in rat. BPC 157 (10 μg, 10 ng, and 10 pg/kg) is given intraperitoneally, once daily; the first application 30 min posttransection, the final 24 h before sacrifice. Throughout the whole 72-day period, BPC 157 consistently improves all muscle-healing parameters (biomechanic, function, macro/microscopy/immunochemistry, finally presentation close to normal non-injured muscle, and no postsurgery leg contracture). Controls exhibit stumps grossly weakly connected (C), at postsurgery day 14 and 28, microscopically (HE, x10), at postsurgery day 14, gap filled with fibrous tissue (white arrow) (c), and at postsurgery day 28, a gap filled with fat tissue (dashed white arrow); incorporating few collagen strands is interposed between the transection stumps and unsuccessful attempts of muscle fibers to cross the gap can be observed (c). Contrarily, BPC 157 rats exhibit stumps grossly well connected, approaching to presentation of the normal non-injured muscle (B), microscopically, at postsurgery day 14 and at postsurgery day 28, broad muscle (black arrow) fibers connecting the stumps, while the fat tissue is much less present (b) (dashed black arrow).

FIGURE 10

Muscle crush injury and BPC 157 therapy (Novinscak et al., 2008; Pevec et al., 2010). Force delivered was of 0.727 Ns/cm² (impulse force 0.4653 Ns, kinetic energy 0.7217 J), to a maximum diameter of gastrocnemius muscle complex (GMC), about 2 cm proximal to the insertion of the Achilles tendon. Regimens with similar effectiveness included (i) BPC 157 dissolved in saline (10 μg, 10 ng/kg body weight), (ii) pentadecapeptide BPC 157 in neutral cream (1.0 or 0.01 μg dissolved in distilled water/g commercial neutral cream). Controls received saline (5.0 ml/kg) applied intraperitoneally and or commercial neutral cream applied as a thin cream layer at the site of injury. All animals were treated only once, immediately after injury, if killed and assessed after 2 h. Alternatively, the animals were treated once daily, receiving a final dose 24 h before death and/or assessment (walking, muscle function, and a macroscopic analysis) at days 1, 2, 4, 7, and 14. Gross posttraumatic hematoma presentation at 2 h after injury induction (C-control (white arrow), B-BPC 157 rats (thin cream layer at the site of the injury immediately after injury induction)). Microscopy assessment (c1, b1 (postinjury day 4), c2, b2 (postinjury day 7), and c3, b3 (postinjury day 14)). HE, x10. Controls. Severe atrophy with severe reduction of myocytes (black arrow) and no regeneration attempt, pronounced perimyocytic edema (postinjury day 4, c1); scarce to moderate regeneratory attempts in muscle with maturing granulation tissue (dashed black arrow) (postinjury day 7, c2); pronounced regeneration with a high number of smaller myocytes and some areas of scarring (postinjury day 14, c3). BPC 157. Clearly visible regenerative activity and less edema (postinjury day 4, b1); florid regenerative activity in myocytes with high number of relatively small myocytes and no scarring (postinjury day 7, b2); well-regenerated myocytes of appropriate size and very little scarring (postinjury day 14, b3).

FIGURE 11

Denervated gracilis muscle and BPC 157 therapy (upper) (Mihovil et al., 2009). Transected muscle induced injured leg function failure as induced leg contracture and BPC 157 therapy (lower) (Staresinic et al., 2006). Presentation of the denervated gracilis muscle (B, C) and normal healthy gracilis muscle (H) in rats at 1 year after denervation. Characteristic denervated muscle presentation in controls (C). Counteraction by BPC 157 (10 μg/kg) therapy given per-orally, in drinking water (0.16 μg/ml, 12 ml/rat/day) till the sacrifice (B). Quadriceps muscle was completely transected transversely 1.0 cm proximal to patella to present a definitive defect that cannot be compensated in rats with a considerable injured leg contracture as presented at postsurgery day 21 in controls with maximal leg extension (c). Counteraction by various regimens of the BPC 157 (10 μg, 10 ng) therapy. Given intraperitoneally, once daily; the first application 30 min posttransection, the final 24 h before sacrifice (b1); per-orally, in drinking water (0.16 μg/ml, 0.16 ng/ml, and 12 ml/rat/day) till the sacrifice (b2); locally, thin layer of neutral cream 1 µg/1 g neutral cream once daily; the first application 30 min post-transection, the final 24 h before sacrifice (b3).

FIGURE 12

(Left, control), (right, BPC 157). Fibrosis and BPC 157 therapy. Summarized gross evidence. Presentation of the bile duct ligation rats (“yellow rats”) following 4 months of occlusion (as a model of the wound-healing response to chronic liver injury), gross rat presentation (C1), and after sacrifice, yellow ears (C2), and liver presentation (C3) (Sever et al., 2019). Possible analogy goes with the dermal, muscle (proximal and distal stump of quadriceps muscle poorly connected at day 72 after transection (c)), tendon (gap after tendon detached from calcaneus (C)) and ligament fibrosis and scar formation, and failed function (Mikus et al., 2001; Krivic et al., 2006; Staresinic et al., 2006; Cerovecki et al., 2010). In rats with bile duct ligation, BPC 157 counteracts cirrhosis and portal hypertension (gross rat presentation (B1), and after sacrifice, normal ears (B2), and liver presentation (B3)) much like it attenuates dermal, muscle (well-formed quadriceps muscle at day 72 after transection (b), tendon (tendon reattached to the calcaneous after detachement (B)), and ligament fibrosis and scar formation, and regained function (Mikus et al., 2001; Krivic et al., 2006; Staresinic et al., 2006; Cerovecki et al., 2010).

FIGURE 13

Healing of the segmental osteoperiosteal bone defect (0.8 cm, in the middle of the left radius) in rabbits (Sebecic et al., 1999). Incompletely healed defect in all controls (assessed during 6 weeks, in 2-weeks intervals (c1 (week 2), c2 (week 4), and c3 (week 6)). Pentadecapeptide BPC 157 beneficial effect was consistently obtained (either percutaneously given locally (10 μg/kg) into the bone defect, or applied intramuscularly (intermittently, at postoperative days 7, 9, 14, and 16 at 10 μg/kg) or continuously (once per day, postoperative days 7–21 at 10 microg or 10 ng/kg)) (b1 (week 2), b2 (week 4), and b3 (week 6)). Comparative regimens included percutaneous administration of autologous bone marrow locally (2 ml, postoperative day 7) as well as an autologous cortical graft inserted in the bone defect immediately after its formation. Saline-treated (2 ml intramuscularly (i.m.) and 2 ml locally into the bone defect), injured animals were used as controls (Sebecic et al., 1999).

FIGURE 14

Corneal ulcer and BPC 157 therapy, ophathalmoscopy and microscopy presentation (Masnec et al., 2015). In deeply anesthetized rats, a penetrant linear 2-mm incision was made and two drops of 0.4% oxybuprocaine topical anesthetic were given (to inhibit possible eyelid reflex) in the paralimbal region of the left cornea at the 5 o’clock position with a 20-gauge MVR incision knife at 45° under an operating microscope. BPC 157 was dissolved in distilled water at 2 pg/ml, 2 ng/ml, and 2 μg/ml, and two eye drops were administered to the left eye in each rat immediately after induction of the injury and then every 8 h up to 120 h; controls received an equal volume of distilled water. Characteristic microscopy presentation (HE, x4) at 48 h after injury induction in control rats was much wider gap, not maturing and abundant granulation tissue, edematous with a lot of fibrin (clot-like) at the surface (white arrow). The surface epithelium is relatively unorganized and progressing over the gap showing a basal cell-like morphology (C) (dashed white arrow). In BPC 157–treated animals, the perforation channel is narrow, partly filled with well-vascularized but maturing, non-edematous granulation tissue (black arrow). The superficial epithelium progressing over the gap looks stratified (B) (dashed black arrow).

FIGURE 15

Total debridement of corneal epithelium and BPC 157 therapy (Lazic et al., 2005). Total debridement of corneal epithelium preformed in rats unilaterally and lesions stained (green) and photographed. Medication was distilled water (control group) or BPC 157 2 pg/ml, 2 ng/ml, and 2 μg/ml, two drops/rat eye started immediately after injury induction, every 8 h up to 40 h (i.e., at 0, 8, 16, 24, 32, and 40 h). Lesions presentation at 24 h in controls (C) and lesions attenuation in BPC 157 rats (B), lesions presentation at 32 h in controls (C) and lesions disappearance in BPC 157 rats (B).