Journal of Investigative Surgery, 26, 141–148, 2013
`Copyright C(cid:2) 2013 Informa Healthcare USA, Inc.
`ISSN: 0894-1939 print / 1521-0553 online
`DOI: 10.3109/08941939.2012.724519
`
`NEW METHODOLOGIES
`
`Comparison of Two Gelatin and Thrombin Combination
`Hemostats in a Porcine Liver Abrasion Model
`Kevin M. Lewis, DVM,1 Holly D. Atlee, DVM,1 Angela J. Mannone,1 Joseph Dwyer, MS,1
`Lawrence Lin, PhD,1 Andreas Goppelt, PhD,2 Heinz Redl, PhD3
`
`1 Baxter Healthcare Corporation, Deerfield, Illinois, USA, 2 Baxter Innovations GmbH, Wien, Austria, 3 Ludwig Boltzmann
`Institute for Experimental and Clinical Traumatology, Auva Research Center Austrian Cluster for Tissue Regeneration,
`Vienna, Austria
`
`ABSTRACT
`
`Background: Surgical hemostasis is achieved using adjunctive hemostats when conventional methods fail.
`Objective: This study compares the effectiveness of two adjunctive gelatin-thrombin hemostats. Hypothesis: To
`determine effectiveness, hemostats were compared in vivo, in vitro, and using scanning electron microscopy
`(SEM). Methods: In vivo, a heparinized porcine liver abrasion model was used to compare hemostatic success,
`degree of bleeding, and blood loss at 2, 5, and 10 minutes post-treatment. In vitro, thrombin in the supernatant
`of each hemostat and Red Blood Cells (RBC’S) in the supernatant of clots formed by each was compared. Results:
`Ultrastructure of one gelatin was smooth and the other stellate. In vivo, smooth gelatin provided superior hemo-
`static success at 5 (85% vs. 60%; OR: 5.3; 95% CI: 1.66 to 17.9) and 10 mins (72.5% vs. 47.5%; OR: 5.0; 95% CI: 1.55 to
`16.1). Smooth gelatin had a statistically different degree of bleeding at 5 (0.58 ± 0.87 [Mean ± SD] vs. 1.03 ± 1.12;
`OR: 3.36; 95% CI: 1.34 to 8.41) and 10 mins (1.13 ± 1.14 vs. 1.65 ± 1.05; OR: 3.87; 95% CI: 1.62 to 9.21). Mean blood
`loss was less with smooth gelatin at 2 (0.07 ± 0.19 vs. 0.13 ± 0.63 ml/min), 5 (0.04 ± 0.13 vs. 0.23 ± 0.45 ml/min),
`and 10 mins (0.09 ± 0.24 vs. 0.21 ± 0.32 ml/min). In vitro, supernatant of smooth gelatin had significantly less
`thrombin (6.81 vs. 10.9 IU/ml, p = .001), and significantly less RBC’s than stellate gelatin (0.07 vs. 0.09 × 106/ul,
`p = .0085). Conclusion: Smooth gelatin has an increased ability to retain thrombin and RBC’s in vitro which may ex-
`plain why it provides superior hemostatic effectiveness, superior control of bleeding, and greater reduced blood
`loss in vivo.
`
`Keywords:
`surgiflo
`
`gelatin; thrombin; hemostat; adjunctive hemostats; hemostatic efficacy; bleeding model; floseal;
`
`INTRODUCTION
`
`Bleeding is an expected complication to all surgi-
`cal procedures. If untreated, bleeding can lead to
`dehiscence, infection, or hematoma resulting in sur-
`gical failure; or lead to hemarthrosis, hemothorax, or
`hemopericardium increasing secondary morbidity and
`mortality. Unlike adhesives and sealants, hemostats are
`intraoperative devices that treat bleeding to increase
`the likelihood of successful surgical outcomes [1–3].
`Topical hemostatic agents include plant- or marine-
`derived material
`(i.e.,
`cellulose, polysaccharides,
`chitosan), nonhuman-derived proteins (i.e., gelatin,
`collagen), human-derived proteins (i.e., thrombin, fib-
`
`rinogen), and/or recombinant proteins (i.e., thrombin,
`aprotinin). Gelatin and topical thrombin are most com-
`monly used, alone or in combination. Gelatin induces
`hemostasis by platelet activation and mechanical
`tamponade, while thrombin cleaves fibrinogen to form
`fibrin [4]. Milled gelatin can be prepared with thrombin
`as a flowable hemostatic agent. Flowable hemostatic
`agents offer unique advantages over nonflowable
`hemostats, such as conforming to wound geometries,
`filling deep lesions, and having the ability to remove
`excess material with irrigation. Flowable hemostatic
`agents are demonstrated to be more effective than
`conventional methods in multiple surgical specialties
`in randomized clinical trials [5–7].
`
`Received 8 March 2012; accepted 22 August 2012.
`Address correspondence to Kevin M. Lewis, Baxter Healthcare Corporation, One Baxter Parkway, Deerfield, Illinois, IL 60015, USA. E-mail:
`kevin lewis@baxter.com
`
`141
`
`ETHICON EXHIBIT 1013
`
`

`

`142 K. M. Lewis et al.
`
`The efficacy of commercially available thrombin de-
`rived from different species and recombinant sources
`have been proven to be similar [8, 9]. The use of human-
`or recombinant-derived thrombin is, however, favored
`because antibodies against bovine thrombin may form
`and attack human Factor V resulting in Factor V de-
`ficiency [7]. In contrast to thrombin, the efficacy of
`commercially available gelatin derived from different
`species has not been investigated though the physio-
`chemical properties of gelatin vary based on the source
`and process of extraction [10].
`This study compared two combination flowable
`hemostats, a bovine gelatin and a porcine gelatin
`hemostat combined with human thrombin, using
`scanning electron microscopy (SEM), and in vivo and
`in vitro test systems. SEM images were used to com-
`pare the ultrastructure of the gelatin size and surface
`variations. An in vivo heparinized porcine liver abra-
`sion model was used to compare in vivo hemostatic
`success, degree of bleeding, and continued blood loss
`after treatment. An in vitro test system was used to
`compare thrombin in the supernatant of each hemostat
`alone, and used to compare the thrombin and the
`number of Red Blood Cells (RBC’s) in the supernatant
`of each hemostat mixed with blood as a metric of clot
`formation.
`
`MATERIALS AND METHODS
`
`Hemostatic Agents
`
`(Baxter
`FLOSEAL VH S/D [Hemostatic Matrix]
`Healthcare Corporation, Deerfield,
`Illinois, USA)
`and SURGIFLO [Hemostatic Matrix] (Ethicon Inc.,
`Somerville, New Jersey, USA) were compared in
`this study. FLOSEAL, containing lyophilized bovine
`gelatin, was prepared with human thrombin provided
`in the hemostatic kit. SURGIFLO, containing partially
`reconstituted porcine gelatin was prepared with
`EVITHROM [Topical Human Thrombin]
`(Ethicon
`Inc., Somerville, New Jersey, USA). All products
`were stored and prepared according to their respec-
`tive Instructions for Use [12–14]. Gelatin products
`were not prepared using the same thrombin, be-
`cause the thrombin from one manufacturer may
`have affected the reconstitution of the other gelatin.
`Collagen procurement, treatment, and other manu-
`facturing processes cannot be controlled for in this
`study.
`FLOSEAL thrombin was reconstituted with calcium
`chloride (40 μmol/ml) for a thrombin concentration
`of approximately 500 IU/ml. FLOSEAL gelatin from a
`10 ml kit was then hydrated with 8 ml of the throm-
`bin solution yielding approximately 10 ml of final
`product with approximately 400 IU/ml of thrombin.
`EVITHROM was thawed and had a thrombin con-
`centration of approximately 1,000 IU/ml. SURGIFLO
`
`gelatin was mixed with 4 ml of thrombin solution yield-
`ing approximately 10 ml of final product with approxi-
`mately 400 IU/ml of thrombin. Both gelatins had the
`same final thrombin concentration of approximately
`400 IU/ml. One lot of each product was used in the
`SEM and in vivo comparisons, while three lots of each
`product were used in the in vitro comparison.
`
`METHODS
`
`SEM
`
`Samples of each hemostat were dehydrated in a
`graded ethanol series to 100% ethanol. They were
`then placed into graded solutions of ethanol and
`hexamethyldisilazane (HMDS)
`followed by fresh
`100% HMDS and allowed to air dry. The specimens
`were then mounted onto aluminum SEM supports
`with carbon tape and coated with palladium for
`conductivity using a Denton Desk IV Sputter/Etch
`Unit (Denton Vacuum, LLC, Moorestown, New Jersey,
`USA). Samples were then morphologically examined
`using a Jeol JSM-7600F Scanning Electron Microscope
`(Jeol USA, Inc., Peabody, Massachusetts, USA) and
`representative images were taken.
`
`In vivo Comparison
`All animal activities were performed according to the
`Animal Welfare Act and The Guide for the Care and
`Use of Laboratory Animals in an AAALAC accredited
`institution. The study protocol was approved by the In-
`stitutional Animal Care and Use Committee prior to
`starting the work. Five female pigs with a mean weight
`of 52.6 kg, ranging 49.2–56.2 kg, were premedicated
`with midazolam (0.3 mg/kg, IM) and mask-induced
`with isoflurane in a 2:1 nitrous oxide-to-oxygen car-
`rier. After intubation, pigs were ventilated and main-
`tained under anesthesia using isoflurane. Warmed lac-
`tated Ringer’s solution was given intravenously at a
`continuous rate infusion throughout the study.
`A heparinized porcine liver abrasion model was
`used to compare the two treatments. This model is a re-
`finement of the liver square model [15]. In this refined
`model, a series of two liver abrasions are created us-
`ing a hand-drill (Dremel Stylus Model 1100-01, Robert
`Bosch Tool Corporation, Mt. Prospect, Illinois, USA)
`fixed with medium grade sandpaper (3M, St. Paul,
`Minnesota, USA). The refined model reduces variabil-
`ity by using standardized 1 cm diameter, 3–4 mm deep
`lesions. The liver abrasion model allows a high num-
`ber of lesions per animal reducing the overall number
`of animals used such that a total of 80 lesions were per-
`formed in a total of 5 animals.
`The treatments were compared at 2, 5, and 10 min
`after application by a single observer, blinded to treat-
`ment. The observer assessed the degree to which each
`
`Journal of Investigative Surgery
`
`

`

`Comparison of Gelatin-Thrombin Hemostats
`
`143
`
`FIGURE 1 Graded scale used to assess the degree of bleeding from each lesion, where “no bleed-
`ing” and “an ooze” are hemostatic success. Each box depicts a treated hepatic abrasion with dif-
`ferent amounts of blood loss.
`
`lesion bleed after treatment using a defined, graded
`scale of 0 to 5; where 0 is no bleeding and 5 is severe
`bleeding (Figure 1) [15]. The degree of bleeding is an
`ordinal metric of hemostasis. While, hemostatic success
`is a binary metric of hemostasis based on the degree of
`bleeding, in which hemostatic success was predefined
`as “No Bleeding” or as an “Ooze.”
`If hemostatic success was not achieved at 2, 5, or
`10 min after treatment, then the rate of blood loss was
`quantified using dry preweighted gauze to collect con-
`tinual blood loss for 1 min. The initial weight of the
`gauze was subtracted from the final weight. The g/min
`were converted to ml/min using a conversion of 1 g
`equaling 1 ml [16]. The rate of blood loss was recorded
`as zero if hemostatic success was achieved because no
`to minimal blood loss could be quantified.
`A research technician randomized the hemostatic
`agents within a lesion series using a random number
`sequence. The same research technician prepared the
`agents in unmarked syringes and presented each to a
`single surgeon who applied the hemostats in the as-
`signed random order at the time of use to avoid test
`item confusion. The surgeon was blinded to the ran-
`domization and hemostatic agent being applied. The
`two lesions in each series were treated at approxi-
`mately the same time to avoid difference in coagulation
`that may result from treating each independently.
`Hemostatic agents were applied to each lesion and
`approximated with saline-dampened gauze for 2 min.
`At 2 and 5 min after application, degree of bleeding
`was assessed and blood loss was measured without
`disturbing the product. Excess product on the two le-
`sions was equally and simultaneously irrigated away
`using saline after the 5-min assessment and blood loss
`measurement. At 10 min after application, bleeding as-
`sessment and blood collection were performed again.
`Differences in coagulation factors and clotting times
`of humans and pigs [17, 18] were overcome by hep-
`arinization. Heparinization also increased the sensitiv-
`ity of the test system by increasing the strength of the
`bleeds. Each pig received a bolus dose of heparin to
`achieve an Activated Clotting Time (ACT) within the
`“safe zone” for cardiopulmonary bypass (i.e., 300 to
`600 s) [19, 20]. The “safe zone” is an objective criteria
`used by perfusionists to determine the safety of using
`cardiopulmonary bypass. The model, therefore, is clini-
`
`C(cid:2) 2013 Informa Healthcare USA, Inc.
`
`cally relevant as it represents appropriate levels of hep-
`arinization mimicking either clinical practice or dis-
`ease. The ACT was measured every 20 min using a clin-
`ical coagulation instrument (Hemochron Whole Blood
`Coagulation System, International Technidyne Corpo-
`ration, Piscataway, New Jersey, USA).
`
`In vitro Comparison
`A thrombin chromogenic assay (DiaPharma Group,
`Inc., West Chester, Ohio, USA) was used to measure
`thrombin in the supernatant of each gelatin when pre-
`pared with thrombin. A 50 μl aliquot of gelatin and
`thrombin, not mixed with blood, was left undisturbed
`◦
`C
`in a tissue culture plate stored in an incubator at 37
`in 5% CO2 for 30 min. Then, 2 mL of phosphate buffer
`solution was added and the sample was stored again in
`◦
`C in 5% CO2 for 30 min. Finally, the
`an incubator at 37
`sample was centrifuged at 2,000 g for 10 min to separate
`the gelatin particles, and the supernatant was collected
`and analyzed.
`A thrombin chromogenic assay (ibid.) and a clin-
`ical hematology instrument (ADVIA R(cid:2)
`2120 Hematol-
`ogy System, Siemens Corporation, New York, USA)
`were used to measure thrombin and RBC’s in the su-
`pernatant of each gelatin when prepared with throm-
`bin and after being mixed with blood, respectively. A
`50 μl aliquot of gelatin and thrombin was mixed with
`300 μl of blood then treated as above. Human Type O
`blood from three separate donors was used. Blood was
`not pooled.
`
`Statistical Analysis
`
`The in vivo sampling unit was the liver lesion with 40
`lesions per group for a total of 80 lesions to detect a dif-
`ference in rates of 75% versus 35%, with an α = 0.05 and
`a 90% power. The statistical analysis was performed on
`the observed degree of bleeding score and hemostatic
`success percent.
`Logistic regression was used to evaluate the treat-
`ment effect at 2, 5, and 10 min post-treatment using
`SAS R(cid:2)
`(SAS Institute Inc., Cary, North Carolina, USA).
`A binomial model of success percent and a propor-
`tional odds model of observed bleeding score were
`used. Independent variables included treatment group,
`
`

`

`144 K. M. Lewis et al.
`
`(A) bovine-derived gelatin
`FIGURE 2 Scanning electron microscopic images of
`(FLOSEAL) with a large-smooth appearance and (B) porcine-derived gelatin (SURGI-
`FLO) with a small-stellate appearance.
`
`pig, liver lobe, and initial bleeding score at baseline.
`The odds ratios and 95% confidence intervals were
`computed at each time point post-treatment. The rate of
`blood loss in ml/min was summarized for comparison.
`The in vitro sampling unit was the supernatant of
`each product with three samples per lot of product per
`blood donor (i.e., a total of 27 samples per product were
`compared). A mixed effect model was used to compare
`mean differences of thrombin and RBC’s before and
`after being mixed with blood. Factors in the model in-
`cluded treatment group, random lot effect nested in
`treatment, random donor effect, treatment by donor in-
`teraction, and donor by lot interaction nested in treat-
`ment. Because of the small number of donors and lots
`in this study, compound symmetry assumption was
`used for the variance-covariance structure.
`
`RESULTS
`
`SEM
`
`The ultrastructure of the two hemostats is drastically
`different (Figure 2). FLOSEAL gelatin is smooth dis-
`crete particles of round and angular shapes, while
`SURGIFLO gelatin is stellate coalescing particles of rib-
`bon shape.
`
`In vivo Comparison
`
`The hemostatic success at 2 min after application
`was similar between smooth gelatin (35 of 40 lesions,
`87.5%) and stellate gelatin (36 of 40 lesions, 90.0%).
`The hemostatic success of the two agents, however, di-
`verged overtime with smooth gelatin providing supe-
`rior hemostatic success to stellate gelatin at 5 min (34 of
`40, 85% vs. 24 of 40, 60%) and 10 min (29 of 40, 72.5% vs.
`19 of 40, 47.5%) after application (Figure 3). The agents
`diverged greatest between 2 and 5 min after application
`and prior to irrigation of excess product. Thereafter,
`the performance was paralleled between the groups
`
`with stellate gelatin having the lower hemostatic suc-
`cess and greater rate of rebleeding over time. The odds
`ratio of binomial success demonstrates the superiority
`of smooth gelatin to stellate gelatin measured at 5 and
`10 min after application (Table 1).
`The mean degree of bleeding at 2 min after appli-
`cation and upon removing the dampened gauze was
`similar between smooth gelatin (0.275 ± 0.679 [Mean
`± SD], N = 40) and stellate gelatin (0.325 ± 0.730, N
`= 40). The degree of bleeding of the two agents then
`diverged between 2 and 5 min after application with
`stellate gelatin yielding a significantly higher degree of
`bleeding (0.575 ± 0.874 vs. 1.025 ± 1.121, N = 40 per
`group). The difference between the agents remained
`similar between 5 and 10 min after application but fa-
`vored smooth gelatin to stellate gelatin (Figure 4). The
`odds ratio of proportional odds demonstrates increas-
`ing superiority of smooth gelatin to stellate gelatin over
`time (Table 1).
`The rate of blood loss prior to treatment was not
`recorded. The rate of blood loss for smooth gelatin
`was 46% lower than stellate gelatin at 2 min (0.07 ±
`
`FIGURE 3 Hemostatic success at each time point post-treatment
`where FLOSEAL, a smooth gelatin, has a hemostatic success per-
`cent much greater than SURGIFLO, a stellate gelatin, over time
`(n = 40 per group per time point). Statistical significance is based
`on an odds ratio of binomial model of success, where FLOSEAL
`with thrombin is significantly different from SURIGFLO with
`thrombin at 5 and 10 min (∗).
`
`Journal of Investigative Surgery
`
`

`

`145
`
`9.2135
`8.4138
`6.2642
`16.0616
`17.7763
`4.4405
`
`1.6216
`1.3394
`0.4786
`1.5489
`1.5950
`0.1926
`
`3.8653
`3.3570
`1.7315
`4.9878
`5.3247
`0.9249
`
`confidencelimit
`
`95%upper
`
`confidencelimit
`
`95%lower
`
`Oddsratio
`
`FLOSEAL/SURGIFLO+Thrombin
`FLOSEAL/SURGIFLO+Thrombin
`FLOSEAL/SURGIFLO+Thrombin
`FLOSEAL/SURGIFLO+Thrombin
`FLOSEAL/SURGIFLO+Thrombin
`FLOSEAL/SURGIFLO+Thrombin
`
`Comparison
`
`.0000
`.0001
`.0000
`.0124
`.0023
`.0004
`
`.3001
`.0131
`.2629
`.2100
`.2691
`.7138
`
`.0000
`.0019
`.5099
`.0176
`.0675
`.8997
`
`5
`2
`
`10
`
`2
`5
`
`10
`
`Hemostaticeffectiveness
`
`–
`–
`
`Controlofbleeding
`
`–
`–
`
`Baselineeffect
`
`p-value
`
`Lobeeffect
`
`p-value
`
`p-value
`Pigeffect
`
`MinutesAfter
`
`application
`
`Model
`
`thevalueabove1.0,thegreaterthesignificance
`oddsratioofproportionalmodelofbleedingscore.A95%lowerconfidencelimitgreaterthan1.0indicatesstatisticalsignificanceandfavorsthenumeratorofthecomparison.Thegreater
`TABLE1Resultsofthemultiplelogisticregressionforhemostaticeffectivenessbasedontheoddsratioofbinomialmodelofsuccesspercent,andforcontrolofbleedingbasedonthe
`
`

`

`146 K. M. Lewis et al.
`
`FIGURE 4 Degree of bleeding at each time point post-treatment
`where FLOSEAL, a smooth gelatin, has the lowest bleeding
`scores over time (n = 40 per group). Statistical significance is
`based on an odds ratio of proportional model of bleeding score,
`where FLOSEAL is significantly different from SURGIFLO with
`thrombin at 5 and 10 min (∗). The whiskers represent plus and
`minus two times the standard error.
`
`0.19 ml/min vs. 0.13 ± 0.63 ml/min, N = 40 per group),
`83% lower at 5 min (0.04 ± 0.13 ml/min vs. 0.23 ±
`0.45 ml/min, N = 40 per group), and 57% lower at
`10 min (0.09 ± 0.24 ml/min vs. 0.21 ± 0.32 ml/min, N =
`40 per group) after application. The range of blood loss
`for stellate gelatin decreased over time, but increased
`on average overtime (Figure 5).
`The mean baseline ACT in this model was 120.8 s
`(N = 5) with a median of 125.0 s and range of 111–128 s.
`The mean heparinized ACT was 498.2 s (N = 71) with a
`median of 407.0 s and a range of 227–1,286 s. The mean
`heparinized ACT was equivalent to 4.2× baseline with
`a median of 3.5× baseline.
`
`In vitro Comparison
`
`Smooth gelatin retained significantly more thrombin
`than stellate gelatin as measured in the supernatant
`of each before being mixed with blood (6.81 vs. 10.89
`IU/ml, p = .0013) and after a clot was formed (0.613 vs.
`1.289 IU/ml, p = .0003) (Table 2). The amount of
`thrombin in the supernatant of each reduced after be-
`ing mixed with blood. Smooth gelatin retained signif-
`icantly more RBC’s than stellate gelatin as measured
`
`FIGURE 5 Box whisker plot for the rate of blood loss 2, 5, and
`10 min after application. FLOSEAL, a smooth gelatin, consis-
`tently had the lowest amount of blood loss over time.
`
`in the supernatant of each after a clot was formed
`(0.0685 vs. 0.0911 × 106/uL, p = .0085).
`
`DISCUSSION
`
`investigated the differences between
`This paper
`two gelatin-thrombin hemostats. The two hemostats
`demonstrated different in vivo performance, which is
`likely due to different ultrastructure. The reason for
`the different ultrastructures was not investigated and
`is of interest for future investigations. The extraction
`method can influence the isoionic point and viscosity
`of the gelatin, which may affect its ultrastructure [11].
`However, the likely difference is that porcine-derived
`gelatin has a higher isoelectric point, lower kinemetic
`viscosity, and different amino acid composition (i.e.,
`decreased alanine, glycine, isoleucine, hydroxyproline,
`and increased tyrosine) than bovine-derived gelatin
`[21]. The species differences are likely the cause for
`the different appearances (i.e. stellate gelatin being
`porcine-derived and smooth gelatin being bovine-
`derived) and subsequent performance differences.
`Smooth gelatin has a superior in vivo performance
`to stellate gelatin over time. The difference may result
`from a greater ability to retain thrombin and ability for
`clot formation, which may be a result of the different
`
`TABLE 2. Mixed Effect Model Results for Mean Difference of Thrombin, Thrombin with Blood, and Red Blood Cells (RBC’s) where,
`statistically, FLOSEAL, containing smooth gelatin, had significantly lower nonretained thrombin and RBC’s than SURGIFLO, containing
`stellate gelatin
`
`Metric
`
`Product thrombin
`(IU/ml)
`Product+Blood
`thrombin (IU/ml)
`RBC’s (×106/ul)
`
`FLOSEAL VH S/D
`mean (n = 27)
`6.81
`
`SURGIFLO+Thrombin
`Mean (n = 27)
`10.89
`
`0.613
`
`0.0685
`
`1.289
`
`0.0911
`
`Mean
`difference
`−4.08
`−0.676
`−0.0226
`
`Lower
`95% CI
`−5.48
`−0.841
`−0.0356
`
`Upper
`95% CI
`−2.67
`−0.511
`−0.0096
`
`p-value of mean
`difference
`
`0.0013
`
`0.0003
`
`0.0085
`
`Journal of Investigative Surgery
`
`

`

`gelatin shapes. The ability to retain a greater thrombin
`concentration in or on gelatin particles increases the
`amount of thrombin delivered to and maintained at
`the site of bleeding over time. While, the ability to
`trap RBC’s indicates that a more effective hemostatic
`plug is formed. The greater thrombin concentration
`and more effective clot formation decreases the time to
`hemostasis, increases control of bleeding, and reduces
`blood loss.
`The lesser ability of stellate gelatin to retain throm-
`bin may explain its inferior performance in vivo. The
`volume of thrombin added to stellate can be increased;
`however, this will likely decrease its viscosity, causing
`more product to be washed away during continued
`bleeding. Alternatively, the concentration of thrombin
`added to stellate gelatin can be increased. But, the con-
`version of fibrinogen to fibrin is maximized at a one
`milligram-to-two unit ratio of fibrinogen to thrombin
`[22, 23]. And, when measured in vivo the natural oc-
`curring concentration of fibrinogen is 145–348 mg/dl
`and concentration of prothrombin is 270–330 U/ml for
`a ratio of 1 mg:122 IU [24, 25]. A thrombin concentra-
`tion greater than 400 IU/ml is then unlikely to increase
`the efficacy of stellate gelatin. Both gelatin forms were
`prepared with the same thrombin concentration, ap-
`proximately 400 IU/ml, in this study.
`The strength of this study is that it used an in vivo
`model relevant to clinical practice, an in vitro model,
`and SEM to offer a possible explanation as to why
`differences are seen. The in vivo differences between
`the products are corroborated by the products’ in-
`dications: FLOSEAL, containing smooth gelatin,
`is
`indicated for all types of bleeding, including arterial
`spurting; SURGIFLO, containing stellate gelatin, is
`indicated for only venous, capillary, and arteriolar
`bleeding when prepared with either thrombin or
`saline [12, 13]. The clinical indications and efficacy
`differences carry clinically meaningful differences.
`The efficacy of the gelatin products was com-
`pared in a retrospective case series of patients under-
`going a laparoscopic partial nephrectomy [26]. Clini-
`cal outcomes—not intraoperative performance—were
`measured in this study. This clinical comparison agrees
`with our data in that a lower amount of blood loss is
`seen with the smooth gelatin (25–650 ml) than with the
`stellate gelatin (50–1,500 ml) [26].
`The limitation of our study is that it only compares
`the agents in one tissue type and in one lesion type,
`a limitation of all standardized bleeding models. This
`limitation, however, is addressed by comparing clini-
`cal data, which represents several different tissue types
`and lesion types. When clinical data is compared, the
`smooth gelatin had a greater percentage of patients
`reaching complete hemostasis at 3, 6, and 10 min after
`application (85%, 93%, 97%, respectively) than with the
`stellate gelatin (64%, 90%, 95%, respectively) [12, 13].
`These clinical differences become more pronounced in
`patients with existing unfavorable clinical conditions
`
`C(cid:2) 2013 Informa Healthcare USA, Inc.
`
`Comparison of Gelatin-Thrombin Hemostats
`
`147
`
`(e.g., hypocoagulopathic conditions due to hepariniza-
`tion, antithrombotic therapies, antiplatelet therapies,
`hemodilution, thrombocytopenia, etc.), as similar to
`our in vivo data.
`
`CONCLUSIONS
`
`This study demonstrates that two flowable gelatin
`hemostats have different ultrastructures which may re-
`sult in in vivo efficacy differences. It was demonstrated
`that smooth gelatin provides superior hemostatic effi-
`cacy, superior control of bleeding, and reduced blood
`loss over time relative to stellate gelatin using a hep-
`arinized porcine bleeding model. This study further
`demonstrated that the smooth gelatin retains signif-
`icantly greater thrombin concentrations and RBC’s
`than stellate gelatin, statistically. The in vitro findings
`may explain the in vivo differences between the two
`gelatins.
`
`ACKNOWLEDGMENTS
`
`The authors thank Jim DiOrio and Mary Ann Mur-
`phy for their imaging expertise and performing the
`SEM, and Huub Kreuwel for reviewing the manuscript.
`The authors also thank the technical and administra-
`tive staff at their respective institutions.
`
`Declaration of Interest: Drs. Kevin M. Lewis, Holly D.
`Atlee, and Lawrence Lin; Mrs. Angela J. Mannone; and
`Mr. Joseph Dwyer are employees of Baxter Healthcare
`Corporation. Dr. Andreas Goppelt is an employee of
`Baxter Innovations GmbH. Dr. Heinz Redl is a consul-
`tant to Baxter Innovations GmbH.
`
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