A study of porcine lung anatomical characteristics and a novel experimental lung transplant model

This study describes some new anatomical findings of the right accessory lobe and a new experimental porcine lung transplant model and our experience with surgical training for the first clinical lung transplant of Vietnam. Subjects and methods: 20 pigs were used for anatomical study. Heart - lung blocks were removed. Methylene blue was selectively injected to different branches of pulmonary artery. Then, all heart - lung blocks were carefully dissected. 30 pigs were used to invent a new surgical model. The caudal lobes of the right lungs were harvested from 15 deceased donors, then transplanted into the opposite position as left caudal lobes (having been rotated 180° along the vertical axis) of 15 recipients following left pneumonectomy. Graft function was determined for 3 hours after reperfusion. Results: Only 13/20 cases, the right accessory lobe veins enter the root of the right caudal lobe veins as described by other authors, 3/20 enter the right inferior pulmonary veins, 2/20 enter the position between the right and the left inferior pulmonary veins, 2/20 enter the left inferior pulmonary veins. Porcine living donor using right lung is impossible, using left lung is possible, but different strategies must be used to preserve the blood supply to the donor’s right accessory lobe. With deceased donor, both the right and left caudal lobes could be used, the right accessory lobes should be excluded. In new surgical model study, all 15 recipients survived, all grafts showed excellent acute and long-term function. Conclusion: A new porcine lobar lung transplant from deceased donor has been invented. This model could be used easily for all anatomical variants.

A STUDY OF PORCINE LUNG ANATOMICAL

CHARACTERISTICS AND A NOVEL EXPERIMENTAL LUNG TRANSPLANT MODEL

Nguyen Trung Chuc 1 ; Do Xuan Hai 1 ; Ngo Thi Dong 1

Thieu Ban Trang 1 ; Trinh Cao Minh 1 ; Do Quyet 1

SUMMARY

Objectives: This study describes some new anatomical findings of the right accessory lobe

and a new experimental porcine lung transplant model and our experience with surgical training

for the first clinical lung transplant of Vietnam Subjects and methods: 20 pigs were used for

anatomical study Heart - lung blocks were removed Methylene blue was selectively injected to

different branches of pulmonary artery Then, all heart - lung blocks were carefully dissected

30 pigs were used to invent a new surgical model The caudal lobes of the right lungs were

harvested from 15 deceased donors, then transplanted into the opposite position as left caudal

lobes (having been rotated 180° along the vertical axis) of 15 recipients following left

pneumonectomy Graft function was determined for 3 hours after reperfusion Results: Only

13/20 cases, the right accessory lobe veins enter the root of the right caudal lobe veins as

described by other authors, 3/20 enter the right inferior pulmonary veins, 2/20 enter the position

between the right and the left inferior pulmonary veins, 2/20 enter the left inferior pulmonary

veins Porcine living donor using right lung is impossible, using left lung is possible, but different

strategies must be used to preserve the blood supply to the donor’s right accessory lobe With

deceased donor, both the right and left caudal lobes could be used, the right accessory lobes

should be excluded In new surgical model study, all 15 recipients survived, all grafts showed

excellent acute and long-term function Conclusion: A new porcine lobar lung transplant from

deceased donor has been invented This model could be used easily for all anatomical variants

* Keywords: Experimental lung transplant; Anatomy characteristics.

INTRODUCTION

Living donor lobar lung transplantation

(LDLLT) is performed as a life-saving

procedure for critically ill patients who are

unlikely to survive the long wait for

cadaveric lungs It has been proved life

saving for various lung diseases and

appears to provide similar or better

survival than cadaveric lung transplantation

On 21 Feb, 2017, Vietnam Military

Medical University has successfully performed performed the first living donor lobar lung transplant in Vietnam The recipient who received a right lower lobe from his father and a left lower lobe from his uncle was a

7 years old boy with end stage idiopathic bronchiectasis diagnosed since birth The donors were his biological father, aged

28 years and his uncle, aged 30 years

1 Vietnam Military Medical University

Corresponding author: Nguyen Trung Chuc (nguyentrungek@gmail.com)

Date received: 20/12/2018

Date accepted: 16/01/2019

This was the result of lung transplantation

program started since 2016 in which an

experimental study was included The

lung transplant in developing countries

such as Vietnam, it is still rare Obstacles

include a lack of experienced surgeons

because we can‟t afford to send doctors

for studying abroad

This study describes some anatomical

characteristics of the right accessory lobe

and a novel porcine experimental lung

transplant model and the implication with

surgical training for lung transplantation

MATERIAL AND METRHODS

The study was done with the approval

of the Animal Experiment and Research

Committee of the institution The animals

were treated according to the Animal

Regulations of the Vietnam Military

Medical University published in 1989

1 Anatomical study

- Methodology: Experimental,

cross-sectioned descriptive study

- Animal preparation: 20 pigs were

used for anatomical study

- Heart - lung block harvest: All pigs

were anesthetized with ketamine (10 mg/kg),

positioned in supine on operation table,

heparinized (100 UI/kg), external jugular

veins and femoral arteries were exposed

and cannulated Normal saline solution

was infused through external jugular vein

catheter and blood was withdrawn from

femoral artery catheter at the same time

until animal death Median stenotomy was

done Heart - lung blocks were removed

with laryngotracheal junction cranially and

aorta piercing the diagphram caudally Methylene blue was selectively injected to different branches of pulmonary artery Then, all heart - lung blocks were carefully dissected by skillfull surgeon using surgical loups

2 New transplant model

- Methodology: Experimental study

- Animal preparation: 30 domestic pigs with the weight from 40 - 50 kg which were divided into 15 donors and 15 recipients were used The weight discrepancy of a donor-recipient pair is not more than

10 kg The animals received no solid food for 18h before the procedure, with water available ad libitum

3 Donor operation

The premedication was done with intramuscular ketamine (10 mg/kg) and atropine sulfate (0.25 mg/kg) During surgery, we placed a peripheral catheter, and previous 100% oxygenation was established as monitored with electrocardiogram (ECG) and pulse-oximetry Anesthesia was induced with propofol (4 mg/kg), fentanyl (3 μg/kg) and atracurium besilate (0.6 mg/kg) through a dorsal vein of the ear Surgical tracheotomy was performed with an endotracheal tube measuring 6 - 7 mm in internal diameter Respiratory assistance included a Dräger SA 1 ventilator Ventilation was controlled by volume (tidal volume 8 mL/kg, 12 - 15 breaths/min, ratio between inspiration and expiration of 1:2) and it was adjusted during surgery in order to maintain in arterial blood between

35 and 40 mmHg of carbonic anhydride;

meanwhile, the inspired fraction of oxygen

(FiO2) was maintained at 1 during the

whole procedure The anesthesia was

maintained with propofol at continuous

perfusion (8 - 10 mg/kg/h), with fentanyl

and atracurium in bolus, as needed The

airway pressure (AWP), blood pressure,

pulmonary arterial pressure (PAP), central

venous pressure, arterial blood gases,

PA blood gases, and cardiac output were

measured The mediastinum was

accessed by a median sternotomy, and

the heart and lungs were exposed by

extended pleural and pericardial incision

The superior and inferior vena cava, the

ascending aorta and the trunk of the

pulmonary artery (PA) were dissected

After administration of heparine (3 mg/kg),

a 18F cannula was inserted into the main

PA through a pursestring suture Previous

to the administration of the flush solution,

250 g of prostaglandin-E1 (alprostadil)

was injected in the PA Then the superior

vena cava was occluded, the aorta

cross-clamped, and the inferior vena cava and

the left atrial appendage were incised for

decompression of the PA flush The lungs

were perfused with 50 - 60 mL/kg of

cold (4°C) low-potassium-dextrin-glucose

solution, at a pressure of 40 cm H2O

Topic cooling of the lungs was performed

by instillation of cold saline solution into

the pleural spaces After the flush,

harvesting of the heart - lung in block was

completed by clamping the trachea at

middle-inspiration

This was followed by the bench surgery,

and the right accessory lobe should be

excluded in advance The accessory lobar

vein, artery and bronchus were dissected

free from surrounding tissue (figure 1)

During the dissection of accessory lobe, care should be taken to avoid possible injuries to the neighbouring structures causing air leak afterwards The accessory lobe vessels were ligated and divided with 4.0 silk suture The bronchus was cut transversely for 4 to 5 mm and sutured with 4.0 polypropylene suture in interrupted fashion After finishing the exclusion of the right accessory lobe, we dissected the pedicle of the right caudal lobe that was going to be reimplanted: the right pulmonary artery (after ligature and sectioning of the cranial and middle lobar branches), the right caudal pulmonary vein (liberated from the pleural adherences until the segmental branches and cut at the root where the right caudal lobe vein enters the left atrium) and the right main bronchus (after cutting and suturing the cranial and middle lobar bronchus)

Figure 1: Exposing pedicle of right

accessory lobe

(Red arrow: The artery; blue arrow: The vein; white arrow: The vein; White

arrow: The bronchus)

Figure 2: Experimental design

(A: The right caudal lobe had been transplanted into the left pleural cavity as left caudal

lobe after having been rotated 180° along the vertical axis; B: The outline illustrating the

implantation of the graft in the left pleural cavity)

4 Recipient operation

The recipient pig was sedated in the

same manner as the donor pig The pig

was placed in the right lateral position,

and a fifth intercostal thoracotomy was

performed The left pulmonary vein, left

main pulmonary artery, and left main

bronchus were seperated, and the left

lung was extracted Heparin (200 U/kg)

was administered intravenously The right

caudal lobe had been transplanted into

the left pleural cavity as left caudal lobe

after having been rotated 180° along the

vertical axis (figure 2A) The pulmonary

structures were anastomosed in the following order: Bronchus, pulmonary artery and pulmonary vein Because the graft was rotated 180° along the vertical axis, everal stratergies was done to prevent the broncho-vascular structures

from tension and kinking (figure 2B)

The garft‟s artery will be posterio-superior

to the graft‟s bronchus, meanwhile the left main pulmonary artery is anterior to the bronchus, so care should be taken during the left pneumonectomy in order to gain the enough lenght of the left main pulmonary artery stump (the left main

pulmonaty artery should be divided distal

to the branch of left apical segment),

in relation to the bronchial structure, the

right lower lobe bronchus (the graft)

should be anastomosed to the left upper

lobe bronchus (recipient) instead of the

left lower lobe bronchus This would

adjust morphological similarity between

the right lower lung lobe with the left

pleural cavity As regards pulmonary

veins, the right lower lobe vein (the graft)

should be connected to the left atrial

appendage instead of the upper

pulmonary vein to avoid excessive strain

Retrograde reperfusion was carried out

first, unclamping the left auricle, followed

by anterograde reperfusion, unclamping

the pulmonar artery Then, the ventilation

of the implanted lobe was performed The

pig was returned to the supine position

The arterial blood gases, PA blood gases,

cardiac output, AWP, PAP, central

venous pressure, and left atrial pressure

were measured at 10, 30, 60, 120, and

180 minutes after reperfusion The right

main pulmonary artery was intermittently

clamped using a tourniquet for 5 minutes

before each measurement The following

3 measured parameters were assigned

to evaluate graft function: Partial pressure

of oxygen in arterial blood gases for

oxygenation or gas exchange; PAP for

problems in vessel anastomosis and

influence of the small pulmonary vascular

beds; and AWP for decreased lung

compliance due to congestion of the graft,

problems with the bronchial anastomosis,

or secretions To investigate the graft

intestinal widening, lung biopsies were taken after 3 hours follow-up Lung tissue samples were inflation-fixed in 10% buffered formalin and stained with hematoxylin and eosin

* Statistical analysis:

The data are expressed as means and standard deviation The statistical study was carried out using the SPSS 20.0 statistical package The ANOVA test was used to detect differences in the evolution

of the variables among the different moments of the experimental procedure The differences were considered statistically significant with a p value of < 0.05

RESULTS

1 Anatomical findings

Only 13/20 cases, the right accessory lobe veins enter the root of the right caudal lobe veins as described by other

authors (fig 3A), 3/20 enter the right inferior pulmonary veins (fig 3B), 2/20

enter the position between the right and

the left inferior pulmonary veins (fig 3C),

2/20 enter the left inferior pulmonary veins

(fig 3D)

2 The new experimental lung transplant model

As for the new lung transplant model, all 15 recipient were survived during follow-up period The transplanted lungs appeared excellent function

To assess lung funcytion, 3 following indicators were selected: Partial arterial oxygen pressure (PaO2), pulmonary artery pressure (PAP) and airway pressure (AWP)

Table 1: PaO2, PAP, AWP values after reperfusion

Pressure of pulmonary artery; AWP (airway pressure): Pressure of airway)

Figure 3: The anatomical findings of effluence of right accessory lobe vein (circle)

(A: Entering the root of the right caudal lobe veins; B: Entering the right inferior

pulmonary veins; C: Entering the position between the right and the left inferior

The differences of values of the PaO2, PAP and AWP indicators at the collecting

data time after reperfusion and baseline values were not statistically significant with

p = 0.2; 0.4 and 0.7, respectively

The macro-image and histological results of 15 samples showed no signs of

interstitial edema (figure 4)

Figure 4: The macro-image and histological results of transplanted lungs after

3 hours of perfusion No evidence of interstitial edema can be found

DISCUSSION

In the anatomical study, the phenotype

of right accessory lobe vein entering the

root of the right caudal lobe veins was

found in 13/20 cases (65%) This result is

slightly different from the result of S

Nikakuki A new finding in our study is the

case of right accessory lobe vein pouring

into the left inferior pulmonary vein (2/20)

Due to the anatomical characteristics

of vascular and bronchial structure, it is

not possible to use right lung for living

donor lung transplant only left lung can be

used Specifically, due to the right apical

segmental bronchus raising directly from

the trachea and the existence of the right

accessory lobe, so it is not likely to

harvest the right lung from living donor as

well as to implant the lung graft into the

right chest cavity In addition, given

detecting the new anatomical phenotype

of the right accessory lobe vein, it is

necessary to take strategy to preserve the

blood supply of the right accessory lobe in

case of this vein entering the left inferior lobe vein while harvesting the left lower lobe in living donor lung transplant model

or performing left pneumonectomy in preparation for lung implantation

In deceased donor lung transplant, only the left lung is usually used as a graft due to the anatomical characteristics mentioned above According to our model created in this study, it is possible to use both right lung and left lung In the case of using the right lung, the right accessory lobe must be cut first To assess lung function after reperfusion, 4 measured parameters were assigned: partial pressure

of oxygen in arterial (PaO2) blood gases for oxygenation or gas exchange; PAP for problems in vessel anastomosis and influence of the small pulmonary vascular beds; and AWP for decreased lung compliance due to congestion of the graft, problems with the bronchial anastomosis,

or secretions and finally histopathological results for microscopic edema In the study of the new lung transplant model,

all 15 recipients survived The graft function

performed well after transplantation and

there were not statistically significant

differences in PaO2, PAP and AWP

between the points of measuring and the

baseline (p > 0 05)

With the new deceased donor lung

transplant model, the use of animal in

researching and training surgical techniques

has been maximized In the old deceased

donor lung transplant model, only left lung

from the donor can be used as the graft

for one recipient Meanwhile, both left and

right lungs from the donor can be used for

2 separate recipients with our model

which can potentially facilitate experimental

lung transplantation and surgical training

In 2015, date reported a successful

lung transplantation case in which the

right lower lobe was transplanted into the

left chest cavity in order to improve size

matching between the donor and the

recipient Our research has deeply

analyzed the differences in the

broncho-vascular system between pigs and people

to invent a new technically feasible lung

transplant model on pigs The

broncho-vascular structure of the right lower lobe

must be carefully evaluated so that when

rotated 180° along the body axis and

implanted into the left chest cavity,

vessels and bronchus of the graft are not

kinked This analysis and evaluation

contributes significantly to the perfection

of surgical techniques as the surgeon

guides the anatomical space in both

normal and abnormal positions

Porcine living donor using right lung is impossible, using left lung is possible, but different strategies must be used to preserve the blood supply to the donor‟s right accessory lobe With deceased donor, both the right and left caudal lobes could be used, the right accessory lobes should be excluded In new surgical model study, all 15 recipients survived, all grafts showed excellent acute and long-term function

CONCLUSION

Lung transplant could be performed successfully in a developing country An creative experiment program, advanced training methods could be substituted for abroad training A new porcine lobar lung transplant from deceased donor has been invented This model could be used easily for all anatomical variants

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