New Approaches to Morphological Diagnosis of Placenta Percreta: Clinical and Morphological Comparisons. Сase Report.

1. Introduction

The term “placenta accreta” was proposed in the last century, when scientists Irving and Hertig first defined it as “abnormal partial or complete attachment of the placenta to the wall of the uterus” [1]. Abnormal attachment and invasion of the placenta villi is manifested in its inability to separate from the wall of the uterus.

As in the FIGO system [2] the proposed subcategories of PAS are designated as grades based on the degree of grossly assessed invasion and local tissue destruction: Grade 1 – Abnormally adherent placenta (pl. accreta) when the villi adhere directly to the myometrium without a decidual interface. Grade 2 – Abnormally invasive placentation (Pl. increta) when the villi invade into the myometrium. Grade 3 – Abnormally invasive placentation (Pl. percreta) when the villi invade the full thickness of the uterine wall either to the serosa or beyond. They are subdivided into: Grade 3a, limited to and including the uterine serosa; Grade 3b, when there is urinary bladder invasion; Grade 3c, when there is invasion of other pelvic tissue. PAS is a dangerous complication of pregnancy leading to maternal morbidity and mortality, associated with a high risk of massive blood loss.

Recently, PAS increased significantly from 1:1200 [3] to 1: 250 - 1:500 in all countries [4,5,6]. Observational studies in United States reported about 1: 272 in 2016 [7]. The most studied and proven factor leading to an increase in the incidence of PAS is number of previous caesarean sections (CS) [2,8,9]. Risk factors for PAS are the previous CS is 3%, 11%, 40%, 61% and 67% for one, two, three, four and five previous CS [5], respectively Uterine scar dehiscence represents an incomplete disruption of the uterine wall at the site of prior cesarean delivery, typically occurs within the anterior lower uterine segment overlying the bladder [10,11].

Risk factors for PAS includes as well the patient’s history of multiple surgical interventions on the uterus (myomectomy, hysteroscopy, curettage), uterine artery embolization, placenta previa and PAS [12], metabolic disorders [13,14,15], which probably initiates inflammation and incomplete regeneration for damaged tissues in the future [4].

2. Сase Report

Patient O*., 37 years old, was admitted to the Pregnancy Pathology Department of the Vidnovsky Perinatal Center in 2022. Singleton pregnancy. Placenta previa. The fetus was in cephalic presentation at 33 weeks and 2 days.

The patient was admitted to Vidnovsky Perinatal Center for therapy aimed at prolonging pregnancy at the period of 33 weeks and 2 days.

Recent pregnancy was the fifth, after the fourth birth. Incompetent scar on the uterus after two CS with the formation of the uterine prolapse or «hernia». Given that ultrasound examination placenta invaded into the uterine scar after previous CS surgery and the wall of bladder, placenta previa was revealed also.

The parity and anamnesis: This is the fifth pregnancy occurred in a natural cycle, no using high reproductive technologies. The first baby was born in 2007 after with spontaneous labor at 38 weeks. A healthy girl was born at 2900 grams. Second pregnancy was in 2012. A healthy boy was born weighing 2690 grams after urgent spontaneous birth. In 2013, the third pregnancy ended in emergency surgical delivery at 38 weeks due to clinically narrow pelvis, a boy was born, (the weight was 4200 g), healthy. The fourth pregnancy was in 2020. There was operative labor at 35 weeks due to severe preeclampsia. The girl was born, 2600 grams, healthy.

The first trimester proceeded without complications. When conducting a screening ultrasound at 13 weeks and 6 days, chorionic presentation on the uterine scar niche was diagnosed. No markers of preeclampsia were identified. Ultrasound examination performed at 19 weeks and 6 days, the myometrium in the area of the uterine scar dehiscence was 1.6 mm. Placenta рrevia was revealed as well. At 33 weeks the pregnant was admitted to the hospital because of elevated risk of premature labor. Ultrasound examination revealed a high risk of PAS as well. Ultrasound examination was performed using transabdominal and transvaginal devises (MEDISON ACCUVIX A30-RUS 2014, Korea and ultrasound diagnostic device of expert class WS80A-RUS 2019, Korea). It was found that the retroplacental myometrium is not visible by ultrasound examination on over 158x133 mm). Aberrant vessels were identified that penetrated to the serosa of the bladder (Figure 1). The placenta was diffusely thickened up to 46 mm, located along the anterior wall, the lower edge of the placenta overlaped the area of the internal os. Ultrasound signs were revealed formation of the uterine prolapse so-called «uterine hernia». High risk of placental invasion into the bladder was detected.

Magnetic resonance imaging (MRI) examination was performed by Siemens MAGNETOM Verio with magnetic field induction 3 T, using standard surface coils. Scanning was performed according to the routine protocol [16,17] with T2-weighted images obtained in three mutually perpendicular planes. А slice thickness of 3-4 mm, field of view of 32-42 cm. T2-weighted images with signal suppression from adipose tissue in the axial plane. T1-weighted images in sagittal and axial planes. T1-weighted images with suppression of MR signal intensity from adipose tissue in any plane, and also with obtaining diffusion-weighted images.

The study performed with a moderately full bladder, when the patient lying on her back. The interpretation of the results was carried out on the basis of the developed unified scoring system for the obtained MAPI data – RADS (Morbidly Adherent Placenta Imaging Reporting and Data System) [18] (Figure 2, Table 1). The developed MAPI-RADS system makes it possible to identify, systematize and quantify the signs of pathological PAS obtained during the study, which makes it possible to establish a class corresponding to the PAS grade and correlating with intraoperative view.

Considering the depth of placental invasion and the high risk of intraoperative massive blood loss, аutoplasma was prepared as the preparation for delivery. Three sessions of donor plasmapheresis were conducted. А result of this 1500 ml of autoplasma were prepared [19].

Planned abdominal delivery was performed at 37 weeks of pregnancy according to author’s technique. [20,21]. Briefly, further improvement of the methodology of organ preserving operations was reflected in the research “Method of operative delivery patients with placenta accretion into the uterine scar"[21]. Briefly, the essence of the technique is to combine tourniquet hemostasis with balloon tamponed of the vagina using a Zhukovsky catheter. The method was presented on Figure 3A-E.

An inferomedial laparotomy was performed with bypass of the umbilicus on the left Сaesarean section was carry out in the area of the uterine fundus. The fetus was removed from the uterine cavity and divided the umbilical cord and immersed in the uterus, the placenta was not separated. Incision on the uterus sewed up in two rows with separate vicryl sutures.

The operation was conducted under long-term epidural anesthesia (at the stage of Cesarean section) with subsequent transition to endotracheal anesthesia (at the stage of metroplasty). After inferomedian laparotomy, the lower uterine segment was examined. Local prolapse of uterine wall (sizes 200x200 mm) with a pronounced vascular network was visualized (Figure 3A).

A caesarean section was performed. A live born full-term boy, with the Apgar score of 7-8 points, was extracted at 6 minutes (the weight =3890 g, the body length =51 cm). After administration of uterotonics there were no signs of placental separation. The umbilical cord was tied and immersed in the uterine cavity. The uterus was with two rows with separate vicryl sutures. Next, the first tourniquet was applied above the «uterine hernia». In the area of the trigonum vesicae (trigonum Lieto). Mobilization was difficult due to placental invasion and severe tissue fibrosis. Below the uterine prolapse, the second uterine tourniquet was placed to capture the bladder in the area of the trigonum vesicae. The «uterine hernia» with an area of placental invasion was excised. The posterior wall of the bladder was resected over the length of 60 mm (Figure 3 B). The uterus with stitches in two rows with separate vicryl sutures. The bladder was sutured in two rows with a continuous vicryl suture. In order to reduce the volume of blood loss, vaginal balloon tamponade was performed using Zhukovsky vaginal catheter.

Intraoperative blood loss was 1547 ml. Two doses of autoplasma equivalent as1000 ml were transfused. 1281 ml of blood were collected intraoperatively, followed by reinfused autologous red blood cells in a volume of 336 ml. The postoperative period proceeded without complications. Postoperative period lasted 7 days.

Placenta and myometrium for morphological research was obtained after Caesarian section. Fragments of the myometrium in the uteroplacental region in PAS zone were excised according to the surgical method researched at the Vidnovsky Perinatal Center [21].

Gross examination: placental weigh was 420 g, dimensions corresponded 18x14x2.5 cm. At one of the edge was revealed area of grayish-red tissue (looks like myometrium) dimensions were 60x40 mm, attached to the maternal surface of the placenta. Thickness this segment varied from 15 mm with thinning up to serous layer (less than 1 mm). Adjacent to this fragment was a grayish area similar to the mucous membrane (of the bladder).

Histological examination. For histological examination, tissue samples were fixed with 10% buffered formalin (#60-001/S, LLC «BioVitrum» Russia). After 24 hours of fixation, the material was washed, dehydrated, and embedded in Histomix Extra paraffin (#10342, LLC «BioVitrum», Russia). Sections from paraffin blocks with a thickness of 4 μm were obtained on a Sakura rotary microtome Accu-Cut SRM200 (Sakura, Japan), stained with hematoxylin and eosin (#07-006, LLC «BioVitrum», Russia), Mallory trichrome kit (#21-036, LLC «BioVitrum», Russia). Microsections placed on glass slides were deparaffinized and re-hydrated in a graded ethanol series, then washed in water and stained with hematoxylin and eosin (LLC «BioVitrum», article: 07-006) or Mallory trichrome kit (#21-036, LLC «BioVitrum», Russia) for detection of connective tissue according to Mallory (LLC «BioVitrum», article: 21-036), then next they were dehydrated and placed in Vitrogel (LLC «BioVitrum», article 12-005) for further microscopic examination.

Immunohistochemical study. Sections from the paraffin blocks were mounted on lysine-coated glass slides (Menzel-Glaser Polysine®), rehydrated, followed by heat-mediated antigen retrieval in citrate solution (pH 6.0), blocked [1 h at RT in 10% goat serum + 0.1% Tween-20 in Tris-buffered saline (TBS)] and incubated overnight at 4˚C with primary antibodies specifically interacting with the antigen on the section, then washed using phosphate buffer. The products of interaction primary antibodies with the antigen were revealed using the horseradish peroxidase conjugate specifically bound to secondary anti-species antibodies. Novolink ™ Polymer Reagent Kit was used to detect bound primary antibodies (#RE7150, Leica, UK) followed by counter-staining with Mayer’s hematoxylin solution (LLC «BioVitrum», article: 05-002/S), dehydration in a graded ethanol series, and mounting with Vitrogel (LLC «BioVitrum», article 12-005). An aqueous solution of 3,3-diaminobenzidine tetrahydrochloride (DAB) was used to stain the product of the immunohistochemical reactions. Products of the positive immunohistochemical reaction were determined as brown staining of the membrane and cytoplasm of cells. For the negative control, the sections were subjected to standard immunohistochemical procedure without incubation with primary antibodies. The positive control was selected according to the specifications manufacturer.

We used mouse monoclonal antibody to cytokeratin 8 (cat# DB098-RTU, DB Biotech) for the detection of epithelial cells. Specimen microscopy was performed using Leica microscope system, consisting of Leica DM2500 microscope, Leica DFC290 video camera image microscopy and analysis software, Image Scope M (Leica, Germany) as well.

All procedures performed in this study involving all patients comply with the ethical standards of the institutions ethics committee and the 1964 Declaration of Helsinki and its subsequent amendments or comparable Ethical standards. This research was approved by the Bioethics Committee at FSBI (protocol No 35 (11) dated 03/23/2022). All included women gave voluntary Agreement to participate in the study, in accordance with the Declaration of Helsinki on the conduct of biomedical research.

As the results of the histological examinations, thinning of the lower segment to a serose membrane with invaded villi presented by an uneven layer of borderline fibrinoid. In myometrium of uteroplacental region multiple villi with dystrophic changes and necrosis, including loss of basophilia of nuclei coated in fibrinoid (“villi - shadows”), with cytotrophoblast remnants were revealed (Figure 4 and Figure 5). The placental villous tree was represented by intermediate villi, angiogenesis with predominantly of branching vessels with unevenly expressed blood supply. Decidual lamina with vast deposits of fibrinoid, and few decidual cells (Figure 4).

In the described structures, moderate lymphoid infiltration with the formation of nodules and vasculitis (chronic cystitis signs). Small foci of squamous metaplasia of the transitional epithelium were observed. Separated muscle bundles and edematous serous membrane with local microvascular thrombosis and/or adjacent retroplacental hematoma zone were detected. There was a large retrochorial hematoma several days old, reaching serous uterine membrane (Figure 4 and Figure 5).

By immunohistochemical examination studies with primary antibody to cytokeratin-8, invasive cytotrophoblast cells were detected in the uterine wall located up to the subserosal layer, including those reaching the adventitia of the walls of subserosal vessels. In the same area, multinucleated giant cells were detected (Figure 5). Multinucleated giant cells were associated to invasion arrested, because in cases of physiological pregnancy they were located at the border of invasion [22].

Moreover, large deposits of fibrinoid were detected in the uteroplacental region. It that fibrinoid is known to be combined product created as a result of both coagulation of plasma proteins and trophoblastic secretion. [23]. We detected increased fibrinoid amount, multiple hemorrhages, areas of dystrophy and necrosis of the myometrium in the uteroplacental region, which significantly disrupted the usual structure of the myometrium, which made the diagnosis unclear.

3. Conclusions

How to interpret these changes according to the latest FIGO classification? So performed study raises a number of questions. How to interpret these changes according to the latest FIGO classification? According to ultrasound and intraoperative data, we show close interactions between the wall of the uterus and the bladder. Assessment of the placental villi invasion depth do not confirm their presence in the bladder wall or parametrium. The formation of adhesions between the placenta and the uterine wall and bladder may be one of the options for PP.

Perhaps, the wall of the bladder is attached to the wall of the uterus through the proliferation of connective tissue, which may be regarded as a variant of pelvic adhesive disease. Should these lesions be classified as a controversial form PP?

Although histological examination considered to be the «gold standard» for verifying the diagnosis, when studying patients with PAS, an integrated approach should be used, taking into account data from visual diagnostic methods (ultrasound, MRI) and intraoperative view. Our study presented a new comprehensive approach to the morphological verification of PP. A detailed analysis in the same cases is necessary. In the future this may require additions and clarifications to the speculated PAS classification.