Pentadecapeptide BPC 157 as Therapy for Inferior Caval Vein Embolization: Recovery of Sodium Laurate-Post-Embolization Syndrome in Rats

After inferior caval vein embolization therapy, post-embolization syndrome (sodium laurate 10 mg/kg, 0.1 mL into rat inferior caval vein, assessment at 15, 30, 60 min, prime lung lesions, thromboemboli occluding lung vessels), as a severe occlusion/occlusion-like syndrome, might be resolved as a whole by stable gastric pentadecapeptide BPC 157 therapy. At 5 min after laurate injection, stable gastric pentadecapeptide BPC 157 was implemented as therapy (10 µg/kg, 10 ng/kg intraperitoneally or intragastrically). As before, confronted with the occlusion of major vessel(s) or similar noxious procedures, such as rapidly acting Virchow triad circumstances, the particular effect of the therapy (i.e., collateral pathways activation, “bypassing vascular key”, i.e., direct blood flow delivery via activation of azygos vein) assisted in the recovery of the vessel/s and counteracted multiorgan failure due to occlusion/occlusion-like syndrome as a whole in the laurate-injected rats. Along with prime lung lesions and thromboemboli occluding lung vessels, post-embolization syndrome rapidly occurred peripherally and centrally as a shared multiorgan and vessel failure, brain, heart, lung, liver, kidney, and gastrointestinal tract lesions, venous hypertension (intracranial (superior sagittal sinus), portal, and caval), aortal hypotension, progressing thrombosis in veins and arteries and stasis, congested and/or failed major veins, and severe ECG disturbances. Whatever the cause, these were all counteracted, eliminated, or attenuated by the application of BPC 157 therapy. As recovery with BPC 157 therapy commonly and rapidly occurred, reversing the collapsed azygos vein to the rescuing collateral pathway might initiate rapid direct blood delivery and start blood flow reorganization. In conclusion, we suggest BPC 157 therapy to resolve further vascular and embolization injuries.

As a highlight (i.e., counteraction of prime lung lesion and thromboemboli occluding lung vessels), all of the BPC 157-treated rats presented rapid and then sustained activation of the azygos vein, enabling direct blood delivery that might help to instantly break the injurious circle.As the common key finding, this is likely to be responsible for the prompt recovery effect.The confirmative proof of the concept appeared as its counteraction mechanism, with the adjacent adverse occlusion/occlusion-like syndrome as a whole being attenuated/eliminated.Although obtained in more demanding noxious conditions, the BPC 157 therapeutic effect was similar to the previous BPC 157 therapy of the mentioned occlusion/occlusion-like syndromes [8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24].

Blood Pressure Disturbances
With BPC 157 therapy, after laurate application, the prompt reduction of the blood pressure disturbances may show a cause-consequence relation, the beneficial effect going on peripherally (portal and caval hypertension and aortal hypotension were almost annihilated) as well as even more centrally (superior sagittal sinus hypertension was attenuated) (Table 1).Otherwise, if therapy was not given, the portal, caval, and even intracranial (superior sagittal sinus) hypertension, as well as the aortal hypotension, rapidly appeared and remained sustainably present until the end of the experiments.

Thrombosis
As the cause-consequence proof of the therapy's effectiveness, thrombosis was promptly reduced by BPC 157 administration (gross assessment), peripherally and centrally (Table 1, Figures 1 and 2).Otherwise, the noxious course was overwhelming, and widespread thrombosis progressed in veins and arteries (i.e., portal, caval vein, superior sagittal sinus, abdominal aorta).
This was along with microscopic thrombus assessment.Analyzing large retroperitoneal vessels, vascular thromboemboli were found in the control rats within the lumen of the abdominal aorta and inferior vena cava in all three assessment time periods (15 min, 30 min, and 60 min) (Figure 1).Thromboemboli consist of fibrin and blood cells.In lung tissue, control rats presented thromboemboli within the lumen of medium and small lung and vascular vessels causing luminal occlusion (Figure 2).They were found in two assessment time periods (30 min and 60 min following application), with regional intra-alveolar hemorrhagic features of lung parenchyma following thromboembolism after the time period of 60 min.Commonly, thromboemboli were absent in BPC 157-treated rats (Fisher exact probability test p < 0.05, at least vs. control).In control rats (small italic letters), thromboemboli (black arrow) were found within the lumen of medium and small lung and vascular vessels causing luminal occlusion, at two assessment time periods (30 min (a) and 60 min (c) following laurate application into inferior caval vein), with regional intraalveolar hemorrhagia (marked area) of lung parenchyma following thromboembolism after the time period of 60 min (e).Thromboemboli were absent in BPC 157-treated rats (capital italic letters) (30 min (B) and 60 min (D) following laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Collateral Pathways, Blood Vessels, and Brain Gross Presentation
Without therapy, post-embolization syndrome occurs with vessel congestion (superior mesenteric vein and inferior caval vein, due to the trapped volume, congested liver and lung), dilated heart, and collapsed vessels (not-functioning azygos vein) and swollen brain) (Figures 3-5).
Without therapy, all of the laurate-injected rats converged to similar effects of continuous vascular failure, and recovery was not able to be spontaneously activated (Figures 3 and 4).The failed collateral pathways presented in each of the given time points failed throughout the complete experimental period.Contrarily, advanced collateral pathways presentation consistently occurred with BPC 157 therapy (either intraperitoneal or intragastric) (Figures 3 and 4).The particular vessel recruitment as a resolution for major vessel failure and stasis counteraction, peripherally and centrally, accords with blood pressure disturbances being attenuated/eliminated, and thrombosis being almost annihilated in In control rats (small italic letters), thromboemboli (black arrow) were found within the lumen of medium and small lung and vascular vessels causing luminal occlusion, at two assessment time periods (30 min (a) and 60 min (c) following laurate application into inferior caval vein), with regional intraalveolar hemorrhagia (marked area) of lung parenchyma following thromboembolism after the time period of 60 min (e).Thromboemboli were absent in BPC 157-treated rats (capital italic letters) (30 min (B) and 60 min (D) following laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).
Only rarely were thromboemboli in the inferior vena cava found in BPC 157-treated rats, and most of these rats had no thromboemboli within the inferior vena cava at all (Fisher exact probability test * p < 0.05, at least vs. control), while no thrombi were found in the abdominal aorta (Figure 1).

Collateral Pathways, Blood Vessels, and Brain Gross Presentation
Without therapy, post-embolization syndrome occurs with vessel congestion (superior mesenteric vein and inferior caval vein, due to the trapped volume, congested liver and lung), dilated heart, and collapsed vessels (not-functioning azygos vein) and swollen brain) (Figures 3-5).Likewise, we posit that this rescuing response results in the counteraction of the increased relative volume of the superior mesenteric vein and inferior caval vein (congestion), which were counteracted and reversed to normal vessel presentation (Figure 4).The consistent outcome is that the presentation of these vessels (Figures 3 and 4) a the heart (Figure 5) returns to close to normal vessel and heart presentation and close normal functioning to re-establish blood flow (multiorgan lesions are largely attenuate due to BPC 157 therapy.As further support, BPC 157 therapy induced a considera change toward normal brain presentation and negative pressure values (Tables 1 and  (i.e., brain swelling occurred with the increased intracranial (superior sagittal sinus) h pertension and increased volume (associated with considerable brain injuries) due to t laurate injection, which was reversed by BPC 157 (Figure 5).Without therapy, all of the laurate-injected rats converged to similar effects of continuous vascular failure, and recovery was not able to be spontaneously activated (Figures 3 and 4).The failed collateral pathways presented in each of the given time points failed throughout the complete experimental period.Contrarily, advanced collateral pathways presentation consistently occurred with BPC 157 therapy (either intraperitoneal or intragastric) (Figures 3 and 4).The particular vessel recruitment as a resolution for major vessel failure and stasis counteraction, peripherally and centrally, accords with blood pressure disturbances being attenuated/eliminated, and thrombosis being almost annihilated in veins and arteries, peripherally and centrally (Table 1).Thus, after laurate inferior caval vein embolization, BPC 157 therapy may fully reverse post-embolization syndrome.in rats that in rats that received 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein, and then saline (control, small italic letters) or BPC 157 therapy (capital italic letters).Presentation immediately after sacrifice (brain) or immediately before sacrifice (heart) at 15 min (a,B,g,H), 30 min (c,D,i,J) and 60 min (e,F,k,L) following application of 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein.

Heart and ECG Disturbances
Commonly, the laureate procedure implicates the prolongation of QTc intervals or PQ intervals, severe bradycardias, and therapeutic evidence in all BPC 157-treated rats (counteraction).The prolongation of QTc intervals or PQ intervals was regularly absent while bradycardia was attenuated.This occurred along with a counteraction of myocardial congestion (Table 3).g,H,i,J,k,L) in rats that in rats that received 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein, and then saline (control, small italic letters) or BPC 157 therapy (capital italic letters).Presentation immediately after sacrifice (brain) or immediately before sacrifice (heart) at 15 min (a,B,g,H), 30 min (c,D,i,J) and 60 min (e,F,k,L) following application of 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein.
Consequently, the particular effects of BPC 157 on the relative volume (Table 2) illustrate the activated defensive response as an immediate therapy effect.There was a reversal of the failed volume.Presentation of azygos vein collapse (as well as the abdominal aorta) was reversed and the azygos vein was reactivated (Figure 3).Likewise, there was a reversal of the increased relative volume (i.e., the superior mesenteric vein and inferior caval vein congestion, which BPC 157 might decrease) (Table 2, Figure 4).As an immediate therapeutic effect, this could be illustrated by gross presentation in Figures 3-5.
The essential point of the therapy was to reactivate the azygos vein, and, thereby, the collapsed volume of the azygos vein (as well as the abdominal aorta) was reversed (i.e., volume increased as the azygos vein was reactivated, thereby enabling direct blood delivery to occur) (Figure 3).
Likewise, we posit that this rescuing response results in the counteraction of the increased relative volume of the superior mesenteric vein and inferior caval vein (congestion), which were counteracted and reversed to normal vessel presentation (Figure 4).
The consistent outcome is that the presentation of these vessels (Figures 3 and 4) and the heart (Figure 5) returns to close to normal vessel and heart presentation and close to normal functioning to re-establish blood flow (multiorgan lesions are largely attenuated) due to BPC 157 therapy.As further support, BPC 157 therapy induced a considerable change toward normal brain presentation and negative pressure values (Tables 1 and 2) (i.e., brain swelling occurred with the increased intracranial (superior sagittal sinus) hypertension and increased volume (associated with considerable brain injuries) due to the laurate injection, which was reversed by BPC 157 (Figure 5).

Heart and ECG Disturbances
Commonly, the laureate procedure implicates the prolongation of QTc intervals or PQ intervals, severe bradycardias, and therapeutic evidence in all BPC 157-treated rats (counteraction).The prolongation of QTc intervals or PQ intervals was regularly absent while bradycardia was attenuated.This occurred along with a counteraction of myocardial congestion (Table 3).After regular laurate inferior caval vein embolization, similar considerable organ lesions indicate a failed common clue (i.e., venous hypertension (intracranial (superior sagittal sinus), portal, and caval) and aortal hypotension, progressed thrombosis, failed collateral recruitment, peripherally and centrally) and advanced post-embolization syndrome.Contrarily, it is likely that the reduced severity of lesions by BPC 157 therapy as an activated clue occurred as result of the immediate impact of the activated collateral pathway as part of the cause-consequence therapeutic course to counteract all of these disturbances.

Heart
Histologic evaluation of representative myocardium tissue showed pronounced congestion and dilatation of coronary arteries and their intramyocardial branches up to the subendocardial area in control rats in all three assessment time periods (15 min, 30 min, and 60 min following laurate application).In BPC 157-treated rats, no or only mild congestion was observed (Table 4, Figure 6).

Lung
We noted, in the control rats, the thickening of the alveolar membranes due to capillary congestion, pulmonary edema, and dilatation of larger blood vessels in all three assessment time periods (15 min, 30 min, and 60 min following laurate application).In addition, focal intralveolar hemorrhage was found at 60 min following application.No changes were found in BPC 157-treated rats (Table 4, Figure 7).Histologic evaluation of representative myocardium tissue showed pronounced congestion and dilatation of coronary arteries and their intramyocardial branches up to the subendocardial area in control rats in all three assessment time periods (15 min, 30 min, and 60 min following laurate application).In BPC 157-treated rats, no or only mild congestion was observed (Table 4, Figure 6).

Lung
We noted, in the control rats, the thickening of the alveolar membranes due to capillary congestion, pulmonary edema, and dilatation of larger blood vessels in all three as-

Liver
Pronounced dilatation of sinusoids and branches of the portal vein in portal tracts was found in the liver tissue of control rats in all three assessment time periods (15 min, 30 min, and 60 min following application).No changes were found in BPC 157-treated rats in the first two assessment time periods (15 min, and 30 min), and only mild congestion of liver parenchyma was found at 60 min following application (Table 4, Figure 8).sessment time periods (15 min, 30 min, and 60 min following laurate application).In addition, focal intralveolar hemorrhage was found at 60 min following application.No changes were found in BPC 157-treated rats (Table 4, Figure 7).

Liver
Pronounced dilatation of sinusoids and branches of the portal vein in portal tracts was found in the liver tissue of control rats in all three assessment time periods (15 min, 30 min, and 60 min following application).No changes were found in BPC 157-treated rats in the first two assessment time periods (15 min, and 30 min), and only mild congestion of liver parenchyma was found at 60 min following application (Table 4, Figure 8).

Kidney
Moderate to marked vascular congestion and interstitial edema were found in control rats in all three assessment time periods (15 min, 30 min, and 60 min) following application.In addition, intratubular hyaline casts were found in control rats at 30 min and 60 min following application.No changes were found in BPC 157-treated rats (Table 4, Figure 9).

Kidney
Moderate to marked vascular congestion and interstitial edema were found in control rats in all three assessment time periods (15 min, 30 min, and 60 min) following application.In addition, intratubular hyaline casts were found in control rats at 30 min and 60 min following application.No changes were found in BPC 157-treated rats (Table 4, Figure 9).2.2.6.Stomach, Small Intestine, and Colon Lesions At 15 min, 30 min, and 60 min after laurate application into the inferior caval vein, the continuation of a noxious course without therapy is associated with hemorrhagic stomach lesions, marked congestion of submucosal blood vessels, and moderate dilatation of intramucosal blood vessels in the stomach and small intestinal and colonic wall.These were completely counteracted in BPC 157-treated rats (Table 4).

2.3.
A Perilous Syndrome Occurred Centrally 2.3.1.Brain Lesions, Cerebral and Cerebellar Cortex, Hypothalamus/Thalamus, and Hippocampus Indicatively, as a common clue that might be the cause of failure (i.e., intracranial (superior sagittal sinus), portal, caval hypertension, aortal hypotension, progressed thrombosis, peripherally and centrally, failed collateral recruitment, disturbed ECG presentation, peripheral organs lesion) without therapy, all of the laurate-injected rats converged to display similar brain lesions as well.Moreover, this might be a very rapid effect, as seen with the comparative presentation of the brain before laurate application in naive rats, the immediate severe brain swelling upon laurate application, and rapid counteraction immediately upon BPC 157 therapy application (Figure 10).Since the immediate post-application period, there was observable gross brain swelling (Table 2, Figure 10) and increased intracranial (superior sagittal sinus) hypertension (and portal and caval hypertension and aortal hypotension) (Table 1).Microscopically, severe brain edema and congestion as well as large intracerebral hemorrhage in the frontoparietal area consistently occurred (Table 5, Figures 11-13).Notably, there is a rapid therapeutic effect observed after BPC 157 application.BPC 157 therapy reduced intracranial (superior sagittal sinus) hypertension and aortal hypotension, and eliminated portal and caval hypertension; additionally, BPC 157 therapy counteracted brain swelling, concurred with only mild brain edema and congestion, lesser brain hemorrhage, and no intraventricular hemorrhage.Indicatively, as a common clue that might be the cause of failure (i.e., intracranial (superior sagittal sinus), portal, caval hypertension, aortal hypotension, progressed thrombosis, peripherally and centrally, failed collateral recruitment, disturbed ECG presentation, peripheral organs lesion) without therapy, all of the laurate-injected rats converged to display similar brain lesions as well.Moreover, this might be a very rapid effect, as seen with the comparative presentation of the brain before laurate application in naive rats, the immediate severe brain swelling upon laurate application, and rapid counteraction immediately upon BPC 157 therapy application (Figure 10).Since the immediate post-application period, there was observable gross brain swelling (Table 2, Figure 10) and increased intracranial (superior sagittal sinus) hypertension (and portal and caval hypertension and aortal hypotension) (Table 1).Microscopically, severe brain edema and congestion as well as large intracerebral hemorrhage in the frontoparietal area consistently occurred (Table 5, Figures 11-13).Notably, there is a rapid therapeutic effect observed after BPC 157 application.BPC 157 therapy reduced intracranial (superior sagittal sinus) hypertension and aortal hypotension, and eliminated portal and caval hypertension; additionally, BPC 157 therapy counteracted brain swelling, concurred with only mild brain edema and congestion, lesser brain hemorrhage, and no intraventricular hemorrhage.
Figure 10.Gross brain presentation in healthy rats (normal small letters) and then after laureate application into inferior caval vein (italic letters), and subsequent application of the BPC 157 therapy (capital italic letters) or saline (small italic letters) (a-h).Brain presentation in normal healthy rats (a,e), brain swelling presentation immediately upon laurate application (b,f).Then, there was the opposite effect of therapy: decreased brain swelling immediately upon BPC 157 administration (C), decreased brain swelling in BPC 157-treated rats immediately before sacrifice (D), in contrast to further brain swelling immediately upon saline administration (g) and increased brain swelling in Figure 10.Gross brain presentation in healthy rats (normal small letters) and then after laureate application into inferior caval vein (italic letters), and subsequent application of the BPC 157 therapy (capital italic letters) or saline (small italic letters) (a-h).Brain presentation in normal healthy rats (a,e), brain swelling presentation immediately upon laurate application (b,f ).Then, there was the opposite effect of therapy: decreased brain swelling immediately upon BPC 157 administration (C), decreased brain swelling in BPC 157-treated rats immediately before sacrifice (D), in contrast to further brain swelling immediately upon saline administration (g) and increased brain swelling in saline-treated rats immediately before sacrifice (h).A similar presentation was noted with both intragastric and intraperitoneal BPC 157 therapy.In the control rats (small italic letters), a pronounced edema and congestion in the brain tissue were observed.Focal, pronounced, and deep intracerebral hemorrhage involving areas of brain tissue was observed, affecting areas of the neocortex, the corpus callosum, the amygdala, and the striatum in the brain tissue (a) (marked area).Moderate and severe neurodegenerative changes of the central nervous system, such as rare karyopyknotic cells affecting the cerebral (c) and cerebellar (e) cortex, a karyopyknosis and degeneration of the Purkinje cells of the cerebellar cortex, and karyopyknosis of cortical neurons, were observed (black arrows), as well as karyopyknosis of cortical neurons, hypothalamic neurons (g), and pyramidal cells of the hippocampus (i) (black arrows).

Brain Damage
All three assessment time periods (15 min (Table 5, Figure 11), 30 min (Table 5, Figure 12), and 60 min (Table 5, Figure 13) following laurate application showed a pronounced edema and congestion in the brain tissue of the control rats).Pronounced and deep intracerebral hemorrhage involves many brain areas, such as the neocortex, the corpus callosum, the amygdala, and the striatum.Intraventricular hemorrhage was not observed.
In contrast, in the BPC 157-treated rats were less affected, presenting with only mild edema and congestion in the brain tissue and intracerebral hemorrhage only in the superficial layers of the neocortex (15 min (Table 5, Figure 11), 30 min (Table 5, Figure 12), and 60 min (Table 5, Figure 13) following laurate application).No intraventricular hemorrhage was found in treated animals.
After the assessment time period of 15 min, control rats presented mild neurodegenerative changes in the central nervous system; rare karyopyknotic cells affecting the cerebral and cerebellar cortex, a karyopyknosis and degeneration of Purkinje cells of the cerebellar cortex, and karyopyknosis of cortical neurons were observed.In contrast, no neurodegenerative changes of the central nervous system were observed in BPC 157-treated rats (Table 5, Figure 11).
Furthermore, brain lesions rapidly progressed, with control rats severely affected; notably, later than 30 min and 60 min in particular.Moderate and severe neurodegenerative changes were widespread, in the cerebral and cerebellar cortex, hypothalamus/thalamus, and hippocampus, affecting all four regions (Table 5, Figures 12 and 13).There was karyopyknosis and degeneration of the Purkinje cells of the cerebellar cortex, and karyopyknosis of the cortical neurons and pyramidal cells of the hippocampus, as well as hypothalamic neurons.BPC 157-treated rats were much less affected, presenting with no or only rare karyopyknotic cells in all four regions, the cerebral and cerebellar cortex, hypothalamus/thalamus, and hippocampus.Thus, in laurate-administered rats, BPC 157 therapy attenuated/counteracted all inferior caval vein embolization-induced brain lesions.
Aortal hypotension was ameliorated.Bradycardia was attenuated.After BPC 157 therapy, laurate-injected rats exhibited no prolonged PQ interval and QTc interval.In addition, BPC 157 therapy, given in any of the regimens (µg, ng, intraperitoneal, intragastric) supports these effects.
In the end, these might be taken as an effective upgrade of the cytoprotection maxim endothelium maintenance → epithelium maintenance (for review, see i.e., [6,25,29]) as a powerful cytoprotective agent rapidly acting to recruit collateral pathways.

Animals
Twelve-week-old male Albino Wistar rats with 200 g body weight, bred in-house at the Animal Pharmacology Facility, School of Medicine, Zagreb, Croatia (registered with the Veterinary Directorate (Reg.No: HR-POK-007)), randomly assigned at six rats/group/interval, were used in all experiments.Rats were acclimated for five days and randomly assigned to their respective treatment groups; housed in polycarbonate (PC) cages (identified with dates, number of study, group, dose, number, and sex of each animal) at 20-24 • C, relative humidity of 40-70%, noise level 60 dB, 12 h of illumination per day (fluorescent lighting), and standard good laboratory practice (GLP) diet and fresh water ad libitum.Procedures were consistent with the standard operating procedures (SOPs) of the Animal Pharmacology Facility and the European Convention for the Protection of Vertebrate Animals used for Experimental and other Scientific Purposes (ETS 123).This study was approved by the local Ethics Committee.Ethical principles of the study complied with the European Directive 010/63/E, the Law on Amendments to the Animal Protection Act (Official Gazette 37/13), the Animal Protection Act (Official Gazette 135/06), the Ordinance on the protection of animals used for scientific purposes (Official Gazette 55/13), the Federation of European Laboratory Animal Science Associations (FELASA) recommendations, and the recommendations of the Ethics Committee of the School of Medicine, University of Zagreb.The experiments were assessed by observers blinded with regards to the treatment.
For assessment at 15 min, 30 min, and 60 min, rats received therapy with BPC 157 (10 µg or 10 ng/kg) or saline (5 mL/kg) (controls) as an early intraperitoneal regimen at 5 min upon laurate administration.For assessment at 15 min, rats received BPC 157 or saline (5 mL/kg) as an intragastric administration at 5 min after laurate injection.

Brain, Heart, Vessel, and Volume Presentation
The applied procedure has been used before in our previous vascular studies [8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24].Brain volume, vessel volume, and heart volume were proportional to the change in the brain, vessel, or heart surface area, respectively.The presentation of the brain and peripheral vessels (superior mesenteric vein, inferior caval vein, azygos vein, and abdominal aorta) was recorded in deeply anesthetized rats, with a camera attached to a VMS-004 Discovery Deluxe USB microscope (Veho, Claymont, DE, USA) [8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24].The border of the brain (or vessels, or heart) in the image was marked using ImageJ software and then the surface area of the brain (or veins, or heart) was measured.This was carried out with brain (or veins, or heart) images for healthy rats, and then for both the control (saline) group and treated (BPC 157) group of rats at the same intervals after the application and at the time of sacrifice.The arithmetic mean of the surface areas was calculated for both groups.Then, the ratio of these two areas was calculated as ( A con A bpc ), where Acon is the arithmetic mean brain (or veins, or heart) area of the control group and Abpc is the arithmetic mean brain (or veins, or heart) area of the treated group.Starting from the square-cube law equations [1] [2], an equation for the change in brain (or veins, or heart) volume proportional to the change in brain (or veins, or heart) surface area [6] was derived.In expressions [1][2][3][4][5], l is defined as any arbitrary one-dimensional length of the brain (for example, the rostro-caudal length of the brain), used only for defining the one-dimensional proportion (l2/l1) between two observed brains (or veins, or hearts) and as an inter-factor (and because of that not measured [6]) for deriving the final expression [6].The procedure was as follows: = × l 2 l 1 2 [1] (square-cube law), = × l 2 l 1 3 [2] (square-cube law), = [3] (from [1], after dividing both sides by A1), = [4] (from [3], after taking the square root of both sides), = [5] (from [2], after dividing both sides by V1), = [6] (after incorporating expression [4] into equation [5]).
We also assessed the neuronal pathological changes in acquired digital images saved as uncompressed 24-bit RGB TIFF files in the software program AnalySIS (Olympus Soft Imaging System GmbH, Munster, Germany), performing quantitative analysis of neuronal damage in the karyopyknotic areas.The neurons of the cortical cerebrum, cerebellar region, hippocampus, and hypothalamus were counted in 10 different high-powered fields (HPF, 400×) and 3 to 5 serial sections of each sample were used to carry out the count as described in [67].The field size was 0.24 µm 2 .
We used four criteria for the estimation of the edema: pale myelin, sieve-like appearance of myelinated areas, dilation of perivascular and pericellular spaces, and vacuolar appearance of the neuropil of gray matter.Edema was graded as heavy, moderate, slight, or no edema (score 0-3) [68].

Statistical Analysis
Statistical analysis was performed by parametric one-way analysis of variance (ANOVA), with the Newman-Keuls post-hoc test or the non-parametric Kruskal-Wallis test and subsequently the Mann-Whitney U test used to compare groups.Values are presented as the mean ± standard deviation (SD) and as the minimum/median/maximum.To compare the frequency difference between groups, the chi-squared test or Fischer's exact test was used.p < 0.05 was considered statistically significant.

Figure 2 .
Figure 2.Thromboemboli presentation in lung (a-e).In control rats (small italic letters), thromboemboli (black arrow) were found within the lumen of medium and small lung and vascular vessels causing luminal occlusion, at two assessment time periods (30 min (a) and 60 min (c) following laurate application into inferior caval vein), with regional intraalveolar hemorrhagia (marked area) of lung parenchyma following thromboembolism after the time period of 60 min (e).Thromboemboli were absent in BPC 157-treated rats (capital italic letters) (30 min (B) and 60 min (D) following laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Figure 2 .
Figure 2.Thromboemboli presentation in lung (a-e).In control rats (small italic letters), thromboemboli (black arrow) were found within the lumen of medium and small lung and vascular vessels causing luminal occlusion, at two assessment time periods (30 min (a) and 60 min (c) following laurate application into inferior caval vein), with regional intraalveolar hemorrhagia (marked area) of lung parenchyma following thromboembolism after the time period of 60 min (e).Thromboemboli were absent in BPC 157-treated rats (capital italic letters) (30 min (B) and 60 min (D) following laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Figure 3 .
Figure 3. Illustrative presentation of azygos vein (AV) (violet arrows) and aorta (AA) in abdomen or thorax (red arrows) and inferior caval vein (blue arrows) (ICV) (a-L) in rats that received saline (control, small italic letters, dashed arrows) or BPC 157 therapy (capital italic letters, full arrows), immediately before sacrifice.Azygos vein and aorta at 15 min (a,B), 30 min (c,D), and 60 min (e,F) following application of 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein.Inferior caval vein and abdominal aorta at 15 min (c,D), 30 min (g,H), and 60 min (k,H) following application of 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein.

Figure 4 .
Figure 4. Illustrative presentation of superior mesenteric vein (SMV, dark blue arrows) and inferior caval vein (ICV, light blue arrows) (a,B,c,D,e,F) in rats that received 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein, and then saline (control, small italic letters, dashed blue arrows) or BPC 157 therapy (capital italic letter, full blue arrows), immediately before sacrifice.Presentation at 15 min (a,B), 30 min (c,D), and 60 min (e,F) following application of 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein.

Pharmaceuticals 2023 , 36 Figure 5 .
Figure 5. Illustrative presentation of brain (swelling) (a,B,c,D,e,F) and heart (dilatation) (g,H,i,J,k,L) in rats that in rats that received 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein, and then saline (control, small italic letters) or BPC 157 therapy (capital italic letters).Presentation immediately after sacrifice (brain) or immediately before sacrifice (heart) at 15 min (a,B,g,H), 30 min (c,D,i,J) and 60 min (e,F,k,L) following application of 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein.

Figure 5 .
Figure 5. Illustrative presentation of brain (swelling) (a,B,c,D,e,F) and heart (dilatation) (g,H,i,J,k,L) in rats that in rats that received 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein, and then saline (control, small italic letters) or BPC 157 therapy (capital italic letters).Presentation immediately after sacrifice (brain) or immediately before sacrifice (heart) at 15 min (a,B,g,H), 30 min (c,D,i,J) and 60 min (e,F,k,L) following application of 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein.

Figure 6 .
Figure 6.Heart, microscopy presentation (a-F).Pronounced congestion and dilatation of coronary arteries and their intramyocardial branches up to the subendocardial area in control rats (small italic letters) at 15 min (a), 30 min (c), and 60 min (e) after laurate application into inferior caval vein.No changes in BPC 157-treated rats (capital italic letters) were observed at 15 min (B), 30 min (D), and 60 min (F) after laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Figure 6 .
Figure 6.Heart, microscopy presentation (a-F).Pronounced congestion and dilatation of coronary arteries and their intramyocardial branches up to the subendocardial area in control rats (small italic letters) at 15 min (a), 30 min (c), and 60 min (e) after laurate application into inferior caval vein.No changes in BPC 157-treated rats (capital italic letters) were observed at 15 min (B), 30 min (D), and 60 min (F) after laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Figure 7 .
Figure 7. Lung, microscopy presentation (a-F).In the control rats, within-lung parenchyma thickening of the alveolar membranes was observed due to capillary congestion, pulmonary edema, and dilatation of larger blood vessels (small italic letters) at 15 min (a), 30 min (c), and 60 min (e) after laurate application into the inferior caval vein.No changes in BPC 157-treated rats were observed (capital italic letters) at 15 min (B), 30 min (D), and 60 min (F) after laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Figure 7 .
Figure 7. Lung, microscopy presentation (a-F).In the control rats, within-lung parenchyma thickening of the alveolar membranes was observed due to capillary congestion, pulmonary edema, and dilatation of larger blood vessels (small italic letters) at 15 min (a), 30 min (c), and 60 min (e) after laurate application into the inferior caval vein.No changes in BPC 157-treated rats were observed (capital italic letters) at 15 min (B), 30 min (D), and 60 min (F) after laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Figure 8 .
Figure 8. Liver, microscopy presentation (a-F).In liver parenchyma, pronounced dilatation of sinusoids and branches of the portal vein in portal tracts was found in liver tissue of control rats (small italic letters) at 15 min (a), 30 min (c), and 60 min (e) after laurate application into the inferior caval vein.No changes in BPC 157-treated rats (capital italic letters) were observed at 15 min (B), 30 min (D), and 60 min (F) after laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Figure 8 .
Figure 8. Liver, microscopy presentation (a-F).In liver parenchyma, pronounced dilatation of sinusoids and branches of the portal vein in portal tracts was found in liver tissue of control rats (small italic letters) at 15 min (a), 30 min (c), and 60 min (e) after laurate application into the inferior caval vein.No changes in BPC 157-treated rats (capital italic letters) were observed at 15 min (B), 30 min (D), and 60 min (F) after laurate application into the inferior caval vein.(HE staining; magnification 400×; scale bar 100 µm).

Figure 11 .
Figure11.Brain neuropathological changes, 15 min following laurate application into the inferior caval vein, (a-J).In the control rats (small italic letters), a pronounced edema and congestion in the

Figure 11 .
Figure11.Brain neuropathological changes, 15 min following laurate application into the inferior caval vein, (a-J).In the control rats (small italic letters), a pronounced edema and congestion in the brain tissue were observed.Focal, pronounced, and deep intracerebral hemorrhage involving areas of brain tissue was observed, affecting areas of the neocortex, the corpus callosum, the amygdala, and the striatum in the brain tissue (a) (marked area).Mild neurodegenerative changes of the central

Figure 12 .
Figure12.Brain neuropathological changes 30 min following laurate application into the inferior caval vein (a-J).In the control rats (italic small letters), a pronounced edema and congestion in the brain tissue were observed.Focal, pronounced, and deep intracerebral hemorrhage involving areas of brain tissue was observed, affecting areas of the neocortex, the corpus callosum, the amygdala, and the striatum in the brain tissue (a) (marked area).Moderate and severe neurodegenerative changes of the central nervous system, such as rare karyopyknotic cells affecting the cerebral (c) and cerebellar (e) cortex, a karyopyknosis and degeneration of the Purkinje cells of the cerebellar cortex, and karyopyknosis of cortical neurons, were observed (black arrows), as well as karyopyknosis of cortical neurons, hypothalamic neurons (g), and pyramidal cells of the hippocampus (i) (black arrows).In the BPC 157-treated rats (capital italic letters), only mild edema and congestion in the brain tissue were observed.Intracerebral hemorrhage was visible only within superficial layers of the neocortex (marked area) (B).BPC 157-treated rats presented no or only rare karyopyknotic cells in all four regions: the cerebrum (D), cerebellum (F), hypothalamus/thalamus (H), and hippocampus (J).(HE staining; magnification 100×; scale bar 200 µm (a,B); magnification 400×; scale bar 100 µm (c-J)).

Figure 12 . 36 Figure 13 .
Figure12.Brain neuropathological changes 30 min following laurate application into the inferior caval vein (a-J).In the control rats (italic small letters), a pronounced edema and congestion in the brain tissue were observed.Focal, pronounced, and deep intracerebral hemorrhage involving areas of brain tissue was observed, affecting areas of the neocortex, the corpus callosum, the amygdala, and the striatum in the brain tissue (a) (marked area).Moderate and severe neurodegenerative changes of the central nervous system, such as rare karyopyknotic cells affecting the cerebral (c) and cerebellar (e) cortex, a karyopyknosis and degeneration of the Purkinje cells of the cerebellar cortex, and karyopyknosis of cortical neurons, were observed (black arrows), as well as karyopyknosis of

Table 3 .
ECG disturbances in rats at 15 min, 30 min, and 60 min following application of 0.1 mL of sodium laurate (10 mg/kg) into inferior caval vein * p ˂ 0.05, at least vs. control.

Table 3 .
ECG disturbances in rats at 15 min, 30 min, and 60 min following application of 0.1 mL of sodium laurate (10 mg/kg) into inferior caval vein * p < 0.05, at least vs. control.

Table 5 .
Lesions were scored microscopically in the cerebrum, cerebellum, hypothalamus, and hippocampus in rats at 15 min, 30 min, and 60 min following the application of 0.1 mL of sodium laurate (10 mg/kg) into the inferior caval vein.* p < 0.05, at least vs. control.