Currently, there is a significant number of patients admitted to surgery departments with trauma of the abdominal cavity's parenchymal organs [1, 2]. This category of patients requires special attention, as their injuries can be complicated by massive intra-abdominal bleeding. Despite the advanced diagnostic equipment available at clinics today, including CT, thromboelastography in specialized hospitals, etc., the proportion of fatal liver and spleen trauma cases remains high, at 20 to 60% [3, 4]. Time is of crucial importance: the quicker the patient receives assistance (counting from the moment of injury), the better are his chances of recovery [5].

In such cases, the key goal of assistance is to stop bleeding, which is achievable not only in the context of a surgery but also with the help of a combination of hemostatics [6]. There are various hemorrhage arrest techniques, from the Pringle manoeuvre through atypical resections to suturing the wound [7]. However, currently, the preferred options are those allowing to preserve organs, enabled by the advancements in electrosurgery (coagulators and high-energy equipment forming the final clot), cryosurgery (non equilibrium plasma or cold plasma), multicomponent polymer sponge implants (MPSI), adhesive compositions (sulfacrylate adhesives), etc. [8]. The latter are gels, sponges, plates, powders; the choice of such product's shape depends on the degree of organ damage and its localization, and the possible pattern of surgery (laparotomy, as a rule, since laparoscopic access is used extremely rarely in urgent situations, with unstable hemodynamics a contraindication thereto) [9].

There are many polymers and organic compounds used as base for such products: gelatin, collagen, cellulose derivatives, etc. The respective MPSIs have proven to be effective, and they are common in clinical practice [10]. The relevance of research in this area is underpinned by a large number of publications by national and foreign authors that cover testing of MPSIs in in vitro and in vivo experiments, the goal of these studies being to find most effective hemostatic that would be highly adhesive and capable of arresting bleeding quickly [11].

This study aimed, in an acute in vivo experiment, to comparatively assess the time and volume of bleeding arrested with new collagen sponge hemostatics combined with Na-CMC.

METHODS

The materials used in this study are the new MPSI ("Composite hemostatic sponge," Russian Federation patent application #2023123284 of September 07, 2023; tab. 1 below lists characteristics thereof), and hemostatics common in clinical practice.

The study was performed on mature male Wistar rats weighing 200–250 g, under general inhalation anesthesia, in two series (liver and spleen) of 60 animals each, divided into 6 groups as per the number of types of tested MPSIs (tab. 1). All surgical interventions were carried out in sterile conditions of the operating unit of the Laboratory of Experimental Surgery and Oncology of the Research Institute of Experimental Medicine of KSMU.

We developed a technique to inflict damage to liver, which included a median laparotomy, liver's left lobe brought out through the wound for marginal resection (10 × 5 × 5 mm) [12]. The injury of the spleen was modeled similarly, with its posterior pole of appropriate dimensions cut off.

The tested sponge, measuring 1.0 × 1.0 cm with a known mass, was applied to the bleeding incision. We registered the volume of blood loss, i.e., how much blood the sponge absorbs, and the time of bleeding. The former (V) was established using the E.M. Levitae gravimetric method, which compares the weight of sterile material before surgery (m1, g) and after (m2, g), when it has siaked up blood. The latter (t, s) was controlled visually and timed with a stopwatch; we lifted the sponge up from the wound every 10 s, and the bleeding was considered arrested when there was no more blood absorbed by it. The animals were removed from the experiment by CO2-induced euthanasia immediately after surgery.

In the context of data processing, we determined the median, 25^th and 75^th percentiles — Me [25;75] (indicators of descriptive statistics). Due to the small size of the sample on the level of groups (n = 10), we established significance of differences with the help of the Mann-Whitney test, and normality of distribution using the Kolmogorov-Smirnov test, with p ≤ 0.05, as acceptable for experimental biomedical research. The software used for the purpose was a licensed version of Statistica 13 Pro (Dell Software Company; Round Rock, USA).

RESULTS

According to the results of series 1 experiments (liver injury), hemorrhage was arrested fastest in group 6, where the new MPSI based on marine collagen and Na-CMC was used. In that group, the bleeding was stopped 2.3 faster than in group 1, where a collagen plate (commonly used in clinical practice) was used (tab. 2, tab. 3). We registered significant differences (twofold and greater) between almost all control groups and group 6, in which the MPSI was 50% collagen, the highest proportion. Group 4, where the MPSI was 15% collagen, also exhibited significant differences with control groups 1 and 2 (sponge plates common in clinical practice).

The bleeding time comparison data given above are supported by the blood loss volume values in the respective study groups (tab. 2, tab. 4). Minimum blood loss was registered in group 6, maximum — in group 3 (MPSI without collagen).

Series 2 (spleen injury) also confirmed efficacy of the sponges developed at KSMU (tab. 2, tab. 5, tab. 6). In group 6, the time of bleeding and the volume of blood loss was at least 1.5 times less than in other test groups.  The former was significantly different between groups 4 and 6 (tab. 5), the latter — significantly different generally (tab. 6).

A noteworthy fact is the lack of differences between new MPSI from group 5 and common hemorrhage arresting products used in control groups. However, the blood loss value registered for the group 5 sample differed from that recorded for group 1. It should also be noted that we have also established significant differences among between test groups (both controlled indicators, series 1 and series 2 experiments).

DISCUSSION

There are numerous published papers that present assessments of MPSI based on collagen and cellulose derivatives (usually, oxidized cellulose) that have already been adopted in clinical practice and currently are a standard for comparison, like Tachocomb and Surgicel Fibrillar. Nevertheless, new MPSI are being intensively developed, because the demand for such products is high, and their users are not satisfied with what is commercially available currently [13, 14]. There are solid philosophies dedicated to the design of such medical commodities, each with a certain opinion regarding their composition. In most cases, foreign manufacturers with established reputation on the market of medical products base their MPSIs on animal collagen or fibers of oxidized and reduced cellulose, medical gelatin, etc. [15, 16].

Authors of this study accumulated data from the experiments designed to assess properties of MPSI based on marine collagen (publications describing it in this capacity are not freely available) and Na-CMC, which is known to prevent commissures, adhere well and have a pronounced hemostatic effect [17, 18].

Considering the acquired data, we can conclude that effectiveness of an MPSI grows together with concentration of collagen therein, which translates into shorter bleeding time and smaller blood loss. Collagen's hemostatic action has been studied sufficiently; it is assumed to trigger coagulation and blood clot formation. The results of our study confirm veracity of this statement for products based on collagen derived from deep-sea squid. Marine collagen has a number of advantages, including low immunogenicity, which reduces the risk of anaphylactic reactions (possible in case of products based on animal collagen), and high hemostatic efficacy that, in a respective MPSI, is boosted by the porous structure of Na-CMC, which adsorbs the liquid component of blood and thus increases concentration of shaped elements in the sponge-injury contact area.

Such products can be made by national manufacturers of medical commodities; they require no expensive imported raw materials. Subsequent studies of these products (reaction of macroorganism tissues, intraoperative and in vitro manipulative properties of MPSI) will allow an assessment of the possibility and prospects of their introduction into the clinical practice of surgical departments.

CONCLUSIONS

The hypothesis tested in this work has the efficacy of MPSI growing due to the addition of collagen (including that of marine origin) to its composition. Based on the resulting data, we can state that the hypothesis was justified: blood loss and bleeding time values were significantly different between control groups and test groups that employed MPSI (six groups, six collagen/Na-CMC ratios). The results of this work are a valid substantiation of further comprehensive testing of the developed MPSIs.