Minimizing bleeding of dorsal venous complex by artificial erection in a rat model
Minimizar la hemorragia del complejo venoso dorsal mediante erección artificial en un modelo de rata

1. Introduction

Santorini’s venous plexus was first described in 1724 by Domenico Santorini [1]. In radical retropubic prostatectomy (RRP) and radical cystoprostatectomy (RCP), one of the most critical steps of the operation is controlling the dorsal venous complex (DVC) due to its high bleeding potential and retropubic location [2]. With the increasing use of laparoscopic and robotic-assisted laparoscopic approaches, DVC control can be achieved more effectively and easily compared to traditional open surgery [3]. However, open RRP and open RCP, which offer similar surgical and functional outcomes, are still widely recommended and practiced in many centers worldwide due to financial and technical limitations [4, 5]. In the literature, several recommendations for controlling DVC have been proposed, including the use of compression after transecting the urethra and DVC, as described by Memmelaar et al. [6]. The ligation of DVC by passing a right-angled clamp below it was described by Chute et al. [7], and the ligation of the DVC using a method described by Reiner and Walsh in 1979 [8]. However, even when attempting to apply these methods, bleeding from DVC can still occur at a high flow rate, complicating the surgical procedure and can increase morbidity.

Penile erection results from increased arterial flow due to vascular changes controlled by the autonomic nervous system, which fills the corpus cavernosum with blood while decreasing venous return through a veno-occlusive mechanism. In this study, considering the physiology of penile erection, we hypothesized that the bleeding from transected DVC could be minimized using a saline-induced artificial erection (AE) technique before transection, thereby reducing the venous return from the penis. We examined whether this bleeding could be controlled by inducing an erection with the AE technique in a rat model with DVC bleeding.

2. Materials and methods

2.1 Surgical procedure

A total of ten healthy male Wistar albino rats, 16 weeks old and weighing between 300 and 350 grams, were utilized. These rats were obtained from the Medipol University Animal Experiments Laboratory in Istanbul, Turkey. The rats were kept at a temperature of 22 ± 1 °C with humidity level around 50 ± 10%. They lived individually ventilated cages that allowed for 10 to 15 ventilations per hour and were maintained on a regular 12-hour night and 12-hour day cycle. The animals had unrestricted access to water and were provided with a standard laboratory diet (Mucedola 4RF21, Milan, Italy). The water was changed, and the cages were cleaned weekly. The cages were lined with aspen chip bedding (Tapvei, Finland). The rats were sedated using intramuscular injections of 4 mg/kg xylazine, and 50 mg/kg ketamine, administered at a body temperature of 37 °C. The subjects were randomly divided into two groups based on whether AE was applied before transection of the DVC: AE+ (n = 5) and AE− (n = 5). The retropubic approach was not considered, as we found that the dorsal vein of rat could be more easily located at the root of the penis, allowing for easier access to the dorsal venous system from this site. In the AE− group, DVC was dissected and transected without AE. Blood from the transected DVC was aspirated, and the total bleeding volume and duration were recorded. The endpoint of the study was defined as a 20% loss of the estimated total volume of blood (4–4.5 mL), calculated as 0.06 × body weight [9]. This amount is approximately equivalent to 750–1000 mL in humans and reflects the average blood loss reported in studies on open RRP [2, 10]. In the AE+ group, AE was initiated by injecting 0.4–0.5 milliliters of sterile saline into the corpus cavernosa using a 28-gauge insulin injector, and then DVC was dissected and transected. Aspiration was used to determine the quantity of blood from the transected DVC. Additional saline injections were administered to maintain it. In two of the five rats, an additional 0.4 mL of saline was given, with the total volume not exceeding 1 mL. On average, 0.6 mL of saline was required per rat to maintain the erection throughout the procedure. The volume and duration of bleeding were recorded. In the AE+ group, aspiration lasted eight minutes, as this was the time required for the AE− group to reach the endpoint. After the procedure, rats were sacrificed by intraperitoneal administration of pentobarbital (R-45Somnopentyl, Kyoritsu Seiyaku Co., Ltd., Tokyo, Japan) at a dose of 200 mg/kg. One penis each from AE+ and AE− groups was excised for histological examination (Fig. 1).

Fig. 1.Surgical dissection of the penis and Deep Dorsal Vein (DDV). (a) Incision line. (b) Dissected penis and DDV. (c) DDV and its proximal bifurcation. (d) Bleeding that occurred in the penis of a rat in the group in which artificial erection (AE) was not applied. (e) Serous fluid coming from the dorsal vein in the penis of a rat in the group in which AE was applied, which shows minimal bleeding. (f) Excised state of the AE−applied penis after the procedure. (g) Excised state of the penis in which AE was not applied.

2.2 Microscopic examination

Rat penile tissues were fixed in 10% formaldehyde, and transverse sections were obtained in successive full-thickness slices. Following standard tissue processing procedures, 4-micron thick sections were taken from the paraffin blocks and the slides were stained with hematoxylin and eosin (Fig. 2).

Fig. 2.Microscopic examination of the penis without (a) and with Artificial Erection (b). (a) Significant congestion in the dorsal vein and other venous structures in the non-erectile tissue (H&E, ×4 magnification). (b) Dilated corpus cavernosum, significantly decreased blood flow and collapse in the dorsal vein and other venous structures in the erectile tissue H&E, ×4 magnification (scale bar = 100 μm) Deep Dorsal Vein (DDV).

3. Results

The duration of aspiration and mean aspirated blood volumes of both groups are given in Table 1. Statistical analysis revealed significant differences in mean blood loss between the two groups during the measurement period.

According to our histological examination, in the AE− group, DVC was normal in size and its lumen was congested. The corpora cavernosa exhibited normal volumes and the lumens of the venous structures around the penis were congested. In contrast, in the AE+ group, the corpora cavernosa were dilated, and the DVC and peripheral vascular structures were collapsed. There was no congestion inside the lumens of the venous structures, and the lumens were observed to be empty.

4. Discussion

In men, prostate cancer is the second most common cancer, while bladder cancer ranks as the seventh most common cancer [11]. At the time of early diagnosis, approximately 25% of bladder cancer patients present with the muscle-invasive stage [12]. Current guidelines strongly recommend RCP in selected cases of T1 and T2 stage bladder cancer cases [13], as well as RP for locally advanced prostate cancer [5]. Therefore, RP and RCP procedures play an important role in urooncology practice. One of the most critical steps common to both surgical procedures is management of the DVC. Even among experienced surgeons, bleeding that requires transfusion can occur during open RP [14, 15].

Since the introduction of Santorini’s venous plexus and the application of radical surgery to the prostate and bladder, various modifications have been proposed to control the DVC. However, a study by Herranz Amo reviewing these methods concluded that there remains no ideal technique to minimize blood loss [16]. In this study, we explored whether bleeding could be reduced using a fundamentally different method. We hypothesized that saline-induced AE, could increase pressure in the subtunical veins could by elevating intracavernous pressure. This, in turn, could decrease blood flow to the dorsal venous plexus by reducing venous return from the corpora cavernosa (Fig. 3).

Fig. 3.Schematic representation of vascular changes in the penis without (a) and with (b) Artificial Erection. (a) Flaccid penile anatomy. (b) Saline injected penile figure, expanded cavernosal sinusoids, and compressed arterial and venous system.

In the AE+ group, clear liquid rather than blood was aspirated from the transected DVC, resulting in less bleeding and a clear view of the surgical field. Furthermore, the volume of aspirated fluid in the AE+ group was significantly lower than the volume of aspirated blood in the AE− group. One reasonable explanation for this decrease is that diminished venous return due to saline-induced AE. Furthermore, the decrease in arterial blood flow to corpora cavernosa, caused by an increase in intracavernosal pressure that exceeds the rat’s systemic blood pressure, may also contribute to the reduction in bleeding. Unfortunately, the blood pressure and pulse values of the rats could not be recorded due to technical limitations. As a further limitation, additionally, could not measure intracavernous pressure during saline injections. However, the volume of saline used to maintain erection aligns with the findings reported by Dai [17]. Despite these limitations, the histological examination revealing the collapse of the DVC and subtunical veins in the AE+ group supports our hypothesis.

5. Conclusions

Our preliminary findings indicate that the AE technique effectively reduces hemorrhage from the transected DVC in rats. While the applicability of these results to human subjects remains uncertain, we believe that this research offers a novel perspective on potential strategies for DVC control during surgical procedures in the future.

Availability of data and materials

The datasets generated and analyzed throughout this study are accessible from the corresponding author upon submission of a reasonable request.

Author contributions

AE and HSG—mainly prepared the study, performed the experiments, and performed the surgery. HSG and AK—write the article. IT—performed the histopathological examination. RS, VC and AI—edited the article. All authors contributed to editorial changes in the manuscript. All authors read and approved the final manuscript.

Ethics approval and consent to participate

This study was conducted following approval from the Medipol University Animal Experiments Local Ethics Committee (approval date: 01 June 2021, reference number: E-38828770-772.02-2456).

Acknowledgment

Not applicable.

Funding

This research received no external funding.

Conflict of interest

The authors declare no conflict of interest.