Ultrasound Guided Erector Spinae Plane Block in Breast Cancer Surgery: Analgesia, Spread and Immunomodulation
Phase: NA
Conditions: Analgesia, Breast Cancer
Sponsor: Alexandria University

 
  
 
ULTRASOUND GUIDED ERECTOR SPINAE PLANE BLOCK 
IN BREAST CANCER SURGERY: ANALGESIA, SPREAD 
AND IMMUNOMODULATION  
23 November 2020 
 
 
Ahmed Mohamed Mohamed Rabah Abdellah 
Assistant Lecturer of Anaesthesia and pain management, 
Medical Research Institute, 
Alexandria University 
Master degree of Anaesthesia and Surgical Intensive Care, Alexndria University, 
2018 
European Diploma of Anaesthesia and Intensive Care, 2018. 
 
 
SUPERVISORS 
 
Prof. Dr. Emad Eldin Abd El Monem Arida 
Professor of Anaesthesia and Surgical Intensive Care, 
Faculty of Medicine, 
University of Alexandria. 
 
Prof. Dr. Nagwa Ahmed Megahed 
Professor of Anaesthesia and Pain Management, 
Medical Research Institute, 
University of Alexandria. 
 
Prof. Dr. Hossam El-Deen Mohammad Ghoneim  

2. 
Professor of Immunology and Allergy, 
Medical Research Institute, 
University of Alexandria. 
 
CO-SUPERVISOR 
Dr. Walid Mohamed Ahmed Mohamed 
Lecturer of Diagnostic Radiology 
Medical Research Institute 
University of Alexandria. 
 
Dr. Wessam Zakaria El-Amrawy 
Lecturer of Anaesthesia and Pain Management, 
Medical Research Institute, 
University of Alexandria. 

3. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

4. 
INTRODUCTION 
Breast cancer is the commonest cancer among Egyptian females (38.8%) 
and modified radical mastectomy (MRM) is the most common surgical 
intervention. (1) 
Breast cancer surgery induces marked acute nociceptive and neuropathic 
pain in the chest wall, axilla and ipsilateral arm.(2) Effective postoperative 
balanced analgesia including regional anaesthesia suppresses the surgical stress 
response and diminishes opioids needs.(3) 
The commonest reasons for overnight stay after breast cancer surgeries are 
postoperative pain, nausea and vomiting and anxiety.(4) Postoperative hospital stay 
for breast cancer patients has shortened owing to not only the less invasive surgical 
technique, but also the efficient pain relief protocols including regional 
anaesthesia.(5) 
Ultrasound‑guided fascial plane blocks are novel techniques which 
showed effectiveness in managing post-mastectomy pain.(6-8) 
Erector spinae plane (ESP) block is one of the fascial plane blocks; its 
efficacy depends on the compartmental spread and the local anaesthetic (LA) 
distribution to nearby target nerves. The absorption and diffusion of the LA has 
a role in determining the quality of ESP block. LA may diffuse anteriorly to the 
ventral and dorsal rami of the spinal nerves and through the intertransverse 
connective tissue to enter the thoracic paravertebral space.(9) 
Local anaesthetic volume and concentration are important factors for 
unilateral ESPB with volumes ranging from 10 to 40 mL have been used.(10) 
However, the optimum volume and concentration, distribution and dermatomal 
coverage is still undetermined.(11)           
As the erector spinae fascia extends from the nuchal fascia cranially to the 
sacrum caudally, local anaesthetic agents extend through several levels, and the 
block can be effective over a large area.(12) 

5. 
Cadaveric studies had been performed to assess the spread of dye in this 
novel block. However, not all of them have had such extensive spread of dye. In 
one study, only minimal spread had been observed into the paravertebral space and 
in another it has been failed to demonstrate any spread at all into the paravertebral 
space or the ventral rami but there was extensive lateral and craniocaudal spread 
around the Erector Spinae (ES) complex. Till now many questions about the extent 
of sensory coverage and if there is difference in the spread between cadavers and 
the living humans, as intrathoracic pressure changes may direct LA spread 
differently.(13, 14) 
Post-mastectomy minimal residual may be unavoidable and the possibility 
for this to result in metastases and cancer recurrence relies on many factors, 
including anti-tumor cell mediated immunity and angiogenic and growth signals 
in sites of the residual.(15) The perioperative neuroendocrine surgical stress 
response, volatile anaesthetics and opioids have deleterious effects on this 
process. In another words; the anaesthetic technique could influence 
perioperative immunosuppression and determine whether the immune response 
would be capable of eradicating the displaced tumour cells.(16-18) 
Natural killer (NK) cells are CD3-- CD56+ lymphocytes playing a pivotal 
role in the innate immune response against cancer. Their main purpose is 
identification and eradication of virus infected cells and metastatic cells. 
Diminished activity of NK cells is a predisposing factor for cancer 
recurrence.(19) Preservation of innate immune function and the direct anti-
inflammatory effects may be the cause of the protective effects of local 
anaesthetics and regional techniques.(20-22) Different concentrations of local 
anaesthetics appear to have different effects on natural killer cells (NKC) 
cytotoxicity. High concentrations of lidocaine suppress NK cell cytotoxicity,(23) 
yet clinically relevant concentrations enhance the in vitro function of NK cells 
via the release of lytic granules.(24) 
 

6. 
AIM OF THE WORK 
The aim of the present study is to evaluate the effect of US guided 
unilateral ESPB using different volumes of local anaesthetic on analgesic 
efficacy, dermatomal spread and immunomodulation in breast cancer surgery. 
The primary outcome is the analgesic efficacy of the different local 
anaesthetic volumes of ESPB. 
The secondary outcomes are the dermatomal dye spread and sensory 
coverage, immunomodulation and complications in breast cancer surgery. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

7. 
PATIENTS 
This study will be carried out on 60 female patients aged between 20-50 
years, admitted to Medical Research Institute Hospital for breast cancer surgery. 
All patients will be of American Society of Anesthesiology (ASA) 
physical status I or II, scheduled for mastectomy under the effect of general 
anaesthesia after taking approval of the Ethical Committee of the Faculty of 
Medicine, and a written consent from every patient included in the study. 
The sample size is determined according to the recommendations of the 
department of biomedical informatics and medical statistics, Medical Research 
Institute using NCSS 2004 & PASS 2000 program. A minimal sample size of 
15 in each group is required to estimate an average difference of 2 in the median 
visual analogue scale (VAS) between groups assuming common standard 
deviation of 2.5, using F test, at level of significance 0.05. 
Patients will be randomly allocated using a computer generated random 
table (Graphpad Software, Inc, La Jolla, CA) and an allocation ratio of 1:1:1. 
Blinding of the research personnel will be maintained throughout the whole 
observation period including all postoperative follow-ups. 
Group (I):  20 patients will receive US-guided deep ESPB block before 
induction of general anaesthesia using 20 ml bupivacaine 0.25 % 
and 5 ml of radiocontrast dye (Omnipaque) at the level of T4. 
Group (II): 20 patients will receive US-guided deep ESPB block before 
induction of general anaesthesia using 40 ml bupivacaine 0.125 % 
and 5 ml of radiocontrast dye (Omnipaque) at the level of T4. 
Group (III): 20 patients will undergo standard general anaesthesia as a control 
group and postoperative analgesia with intravenous morphine 
patient controlled analgesia (PCA) and rescue analgesia if 
required. 

8. 
Exclusion criteria 
1. Age below 18 or more than 70. 
2. Scoliosis or any vertebral anomalies or previous spinal surgeries. 
3. Morbid obesity (BMI ≥ 40 kg/m2). 
4. Allergy or contraindication to any of the studied medications or 
anaesthetic agents. 
5. Chronic opioid analgesic use. 
6. Pregnancy. 
7. Infection at the site of injection or any other contraindication for regional 
anaesthesia. 
8. Duration of surgery more than 90 minutes. 
9. Renal impairment. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

9. 
METHODS 
Preoperative evaluation of the patients: 
Evaluation of the patients will be carried out on the day before surgery 
through proper history taking, clinical examination, complete blood count 
(CBC) and needed laboratory investigations according to the patient status. 
Plain X ray chest and ECG will be done for patients above 40 years. All patients 
will be informed with the procedure of US guided deep ESPB block and they 
will be trained to use the visual analogue scale (VAS) which consists of 10 cm 
line, 0 equivalent to no pain and 10 denoting the worst imaginable pain.(25)  
Preanaesthetic preparation and premedication: 
On arrival to the operative theatre a peripheral venous catheter will be 
inserted to all patients, multichannel monitor (Vamos- Dragar-Germany) will be 
attached to the patient to display ECG (lead II), heart rate (beats/min), non-
invasive mean arterial blood pressure (mmHg) and peripheral oxygen saturation 
(SpO2%). All patients will receive midazolam (0.05mg/kg) and fentanyl 
(0.5μg/kg) intravenously 3 minutes before performance of the block. 
Technique of ultrasound guided deep ESPB block: 
Ultrasound guided deep ESPB block will be performed in the block room 
at Medical Research Institute hospital. Patients will be placed in the prone 
position and a high-frequency linear probe (L 6-12MHz) of SonoSite, S nerve, 2 
D machine, USA will be sterilized. After skin sterilization, the ultrasound probe 
will be placed 2.5–3 cm laterally to the spinous process in a parasagittal oblique 
plane, at the seventh cervical vertebra and moved caudally till T4. 
After anatomical scanning and identification of the transverse process of 
T4 and the three muscles (Trapezius, Rhomboid major and Erector spinae), 2 ml 
of lidocaine 2% will be used to numb the skin then 18-gauge Tuohy needle will 
be advanced cranio-caudally towards the lateral border of transverse process of 
T4 using the in-plane technique. 

10. 
The needle tip will be located in the fascial plane between the transverse 
process and erector spinae muscle. The correct needle position will be tested by 
injecting 2 ml of saline resulting in hydrodissection of the plane followed by the 
injection of: 
o 20 ml bupivacaine 0.25% and 5 ml of radiocontrast dye (Omnipaque) in 
group 1. 
o 40 ml bupivacaine 0.125% and 5 ml of radiocontrast dye (Omnipaque) in 
group 2. 
Then, insertion of epidural catheter 2 to 3 cm over the tip of the Tuohy 
needle under real-time US guidance will be done. 
Assessment of deep ESP sensory block: 
Field of sensory block from T1 to T6 will be assessed bilaterally every 3 
minutes for 15 minutes after deep ESPB using a piece of cotton soaked in iced 
water. The adequacy of sensory block T1-T6 will be determined before 
induction of general anaesthesia. 
Assessment of ES dye spread: 
After deep ESPB, patients will be transferred to the radiology suite where a 
CT scan of the thoracic region will be performed. The spread of the injected 
solution in the deep ESP will be evaluated using DICOM image processing for 
Mac (OsirixX, PixmeoSARL; Bern, Switzerland) and a three-dimensional 
digital reconstruction of the distribution of the injected contrast will be 
obtained. 
Anaesthesia 
All patients will receive standardized anaesthetic technique. General 
anaesthesia will be induced in each group by intravenous fentanyl 1μg/kg, 
propofol (2.5 mg/kg) and cisatracurium (0.15 mg/kg) to facilitate endotracheal 
intubation. Anaesthesia will be maintained with isoflurane (1.2 – 1.5%) and 

11. 
oxygen/air mixture (50 %, 50 %). Incremental doses of cisatracurium 0.03 
mg/kg will be given to maintain anaesthesia guided by train of four (TOF) count 
using the nerve stimulator module of (TOF watch –Organon-Ireland). 
Ventilation will be maintained at a tidal volume of 6 ml/kg and a rate to adjust 
the end-tidal carbon dioxide at (35-40 mmHg) using the ventilator (Fabius GS- 
Drager-Germany). 
At the end of surgery, anaesthesia will be discontinued, residual 
neuromuscular block will be antagonised by atropine 0.01 mg/kg and neostigmine 
0.04 mg/kg, the trachea will be extubated and patients will be transferred to the 
postoperative anaesthesia care unit (PACU) for the next 24 hours. 
Postoperative analgesia: 
Bolus of bupivacaine, with the same volume and concentration assigned to 
each group, will be given in the epidural catheter at the end of surgery in groups I 
& II. 
In the PACU, pain score (as with other measurements) will be assessed by a 
physician not involved in the study design. 
Intravenous morphine patient controlled analgesia (PCA) will be prepared by 
50 mg morphine +45 ml normal saline with a concentration of 1 mg morphine /1 
ml. 
1. Bolus dose of 0.05 mg/kg. 
2. Lockout Interval: 10 minutes. 
3. Four hourly limiting doses will be 10 mg. 
If VAS still ≥ 4, patients will be administered Ketorolac 30 mg IV as rescue 
analgesia. 
Natural killer cells isolation and cytotoxicity assay:(16, 18) 
Samples of 1 ml of patients’ peripheral blood will be collected on EDTA for 
flow cytometry to enumerate for both cytotoxic lymphocyte populations (NK cells 

12. 
and cytotoxic t lymphocytes (Ctls)). CD 56 will be used as a marker for NK cells, 
while CD 8 will be used as a marker for Ctls. 
Cytotoxicity assay will be done by measuring the release of lactate 
dehydrogenase (LDH) from non-viable cells (Cytotoxicity Detection Kit, 630117; 
Clontech 
Laboratories, 
Mountain 
View, 
California) 
according 
to 
the 
manufacturer’s instructions. Then ratio of LDH released specifically from NK cells 
will be correlated according to the results of flow cytometry.(16, 18) 
 
 

13. 
Measurements 
The following measurements will be carried out: 
1. Patient demographics including age, sex and weight. 
2. Vital signs:  
1. Heart rate (beats/min) and ECG will be continuously monitored and will 
be recorded before induction of anaesthesia, every 15 minutes 
intraoperatively and at arrival to the PACU, then every 4 hours for the 
first 24 postoperative hours with recording of any abnormalities. 
2. Peripheral oxygen saturation (SpO2%) will be continuously monitored 
and will be recorded before induction of anaesthesia, every 15 minutes 
intraoperatively and at arrival to the PACU, then every 4 hours for the 
first 24 postoperative hours. 
3. Mean arterial blood pressure (MABP) in mmHg will be monitored and 
recorded before induction of general anaesthesia, every 15 minutes 
intraoperatively and at arrival to the PACU, then every 4 hours for the 
first 24 postoperative hours. 
4. Postoperative respiratory rate will be recorded every hour for the first 4 
hours and then every 4 hours for the rest of the 24 hours. 
3. Patient sedation scale:(28) 
Patient sedation will be assessed by Richmond Agitation-Sedation scale in 
PACU every 4 hours for 24 hours. 
 
 
 
 

14. 
Table (1): Richmond agitation sedation scale(28) 
Score 
Term 
Description 
+4 
Combative 
Overtly combative or violent; immediate danger to 
staff 
+3 
Very agitated 
Pulls on or removes tube(s) or catheter(s) or has 
aggressive behaviour toward staff 
+2 
Agitated 
Frequent nonpurposeful movement or patient–
ventilator dyssynchrony 
+1 
Restless 
Anxious or apprehensive but movements not 
aggressive or vigorous 
0 
Alert and calm 
Spontaneously pays attention to caregiver 
-1 
Drowsy 
Not fully alert, but has sustained (more than 10 
seconds) awakening, with eye contact, to voice 
-2 
Light sedation 
Briefly (less than 10 seconds) awakens with eye 
contact to voice 
-3 
Moderate sedation 
Any movement (but no eye contact) to voice 
-4 
Deep sedation 
No response to voice, but any movement to physical 
stimulation 
-5 
Unarousable 
No response to voice or physical stimulation 
4. Pain assessment: 
1. Acute postoperative nociceptive pain will be measured by: 
a. Visual analogue scale (0-10) where 0=no pain, 10=worst imagineable 
pain every hour for the first 4 hours and then every 4 hours for the 
rest of the 24 postoperative hours. This will be done during rest and 
movement. 
b. Total amount of IV morphine will be calculated at the end of the 24 
postoperative hours. 
c. Total amount of rescue analgesia will be calculated and first time of 
its need will be recorded. 
2. Acute postoperative neuropathic pain will be detected 10 days after 
surgery by DN4 (Douleur Neuropathique 4) questionnaire (10 Questions 
with cut-off value 4/10)(26). 

15. 
 
 
Before patients would be discharged home, DN4 questionnaire will be 
explained to every patient. Postmastectomy neuropathic pain will be detected in 
breast, axilla or arm. Its onset (days) and interference with work, sleep or mood 
will be recorded. 
 
 

16. 
5. Post-block complications: 
Any complications after the block will be spotted and recorded: 
1. Symptoms or signs of local anaesthetic toxicity (tinnitus, perioral 
numbness, seizure). 
2. Postoperative nausea and or vomiting (PONV) score: 
1. None. 
2.  Yes, and relieved by treatment. 
3. Yes, but not relieved by treatment. 
Intravenous metoclopramide (10 mg) and ondansetron (4 mg) will be given 
as the first and second line of treatment of vomiting, respectively. 
3. Arrhythmias. 
4. Haematoma at the site of injection. 
5. Respiratory depression (when respiratory rate < 8 breath / min). 
6. CT scan for the spread: 
Patients will be imaged 15 minutes after the block and before induction of 
general anaesthesia and surgery. 3 D reconstruction of the image will be done 
then Craniocaudal spread of the contrast and spread to Paravertebral space, 
epidural space or rami of the spinal nerves will be assessed and recorded. 
7. Immunomodulatory outcome: (16, 18) 
Cytotoxicity of both lymphocyte populations will be assessed by the 
measuring LDH by enzyme-linked immunosorbent assay or Western blot before 
deep ESPB and 24 hours after the block. 
Then ratio of LDH released specifically from NK cells will be correlated 
according to the results of flow cytometry for both cytotoxic lymphocyte 
populations (NK cells and cytotoxic t lymphocytes (Ctls)). 
 

17. 
8. Patient satisfaction scale:(27) 
Before the patients would be discharged in the second postoperative day, 
their satisfaction with medical care and the method of postoperative pain relief 
will be measured by Likert scale, which is a self-report scale where 0 = strong 
dissatisfaction, 1 = dissatisfaction, 2 = neutral, 3 = satisfaction and 4 = strong 
satisfaction (for each item of the scale). 
Figure (1): Patient satisfaction scale:(27) 
 
 
 
 

18. 
ETHICS OF RESEARCH 
Research on human or human products: 
 
Prospective study: informed consent will be taken from patients. In case of 
incompetent patients the informed consent will be taken from the 
guardians. 
 
Retrospective study: confidentiality of records will be considered. 
 
DNA/genomic material: informed consent for DNA / genomic test and for 
research will be taken from patients. No further test will be carried out 
except with further approval of committee and patients. If the samples will 
travel outside Egypt the researcher will be responsible for transportation 
and security approval. 
 
All drugs used in the research are approved by the Egyptian Ministry of 
Health. 
 
Research on animal: 
 
The animal species are appropriate for the test. 
 
After test, if animal will suffer, it will be euthanized and properly disposed. 
 
After operation, it will have a proper postoperative care.  
 

19. 
 
RESULTS 
Results obtained from this study will be tabulated and statistically analysed 
using standard statistical methods. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

20. 
DISCUSSION 
The results obtained from this study will be discussed in view of 
achievement of the aim and compared with available published data in the same 
field of research. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

21. 
REFRENCES 
1. 
Manzour AF, Gamal Eldin DA. Awareness about breast cancer and 
mammogram among women attending outpatient clinics, Ain Shams 
University Hospitals, Egypt. J Egypt Public Health Assoc. 2019 Dec 
4;94(1):26. 
2. 
Fecho K, Miller Nr, Merritt Sa, Klauber‑Demore N, Hultman Cs, Blau 
Ws. Acute and persistent postoperative pain after breast surgery. Pain Med. 
2009;10:708‑15. 
3. 
Gärtner R, Jensen Mb, Nielsen J, Ewertz M, Kroman N, Kehlet H. 
Prevalence of and factors associated with persistent pain following breast 
cancer surgery. JAMA 2009;302:1985‑92. 
4. 
S. Marla, S. Stallard. Systematic review of day surgery for breast cancer. 
Int J Surg. 2009;7(4):318-23.  
5. 
Arsalani-Zadeh R, Elfadl D, Yassin N, Macfie J. Evidence-based review of 
enhancing postoperative recovery after breast surgery. Br J Surg. 2011; 
98(2):181-96.  
6. 
Singh S, Chowdhary NK. Erector spinae plane block an effective block for 
post‑operative analgesia in modified radical mastectomy. Indian J 
Anaesth. 2018;62:148-50. 
7. 
Veiga M, Costa D, Brazão I. Erector spinae plane block for radical 
mastectomy: A new indication? Rev Esp Anestesiol Reanim (English 
Edition). 2018;65:112-5. 
8. 
Gürkan Y, Aksu C, Kuş A, Yörükoğlu UH, Kılıç CT. Ultrasound guided 
erector spinae plane block reduces postoperative opioid consumption 
following breast surgery: A randomized controlled study. J Clin Anesth. 
2018;50:65‑8. 

22. 
9. 
Pourkashanian A, Narayanan M, Venkataraju Ar. The Erector Spinae 
Plane Block: A Review of Current Evidence. Anaesthesia Tutorial week; 
2019. 
10. Tulgar S, Ahiskalioglu A, De Cassai A, Gurkan Y. Efficacy of bilateral 
erector spinae plane block in the management of pain: current insights. J 
Pain Res. 2019;12 2597–613. 
11. Altıparmak B, Korkmaz Toker M, Uysal Aİ, Turan M, Gümüş Demirbilek 
S. Comparison of the effects of modified pectoral nerve block and erector 
spinae plane block on postoperative opioid consumption and pain scores of 
patients after radical mastectomy surgery: A prospective, randomized, 
controlled trial. J Clin Anesth. 2019;54:61‑5. 
12. Forero M, Rajarathinam M, Adhikary S, Chin KJ. Erector spinae plane 
(ESP) block in the management of post thoracotomy pain syndrome: a case 
series. Scand J Pain. 2017;17:325–9. 
13. Ivanusic J, Konishi Y, Barrington MJ. A cadaveric study investigating the 
mechanism of action of erector spinae blockade. Reg Anesth Pain Med. 
2018;43(6):567-71. 
14. Yang HM, Choi YJ, Kwon HJ, O J, Cho TH, Kim SH. Comparison of 
injectate spread and nerve involvement between retrolaminar and erector 
spinae plane blocks in the thoracic region: a cadaveric study. Anaesthesia. 
2018;73(10):1244-50. 
15. Sessler DI, Ben-Eliyahu S, Mascha EJ, Parat MO, Buggy DJ. Can regional 
analgesia reduce the risk of recurrence after breast cancer? Methodology of 
a multicenter randomized trial. Contemp Clin Trials. 2008; 29(4):517-26.  
16. Buckley A, Mcquaid S, Johnson P, Buggy DJ. Effect of anaesthetic 
technique on the natural killer cell anti-tumour activity of serum from 
women undergoing breast cancer surgery: a pilot study. Br J Anaesth. 
2014; 113 Suppl 1:i:56-62.  

23. 
17. Tazawa K, Koutsogiannaki S, Chamberlain M, Yuki K. The effect of 
different anesthetics on tumor cytotoxicity by natural killer cells. Toxicol 
Lett. 2017; 266:23-31.  
18. Dong H, Zhang Y, Xi H. The effects of epidural anaesthesia and analgesia 
on natural killer cell cytotoxicity and cytokine response in patients with 
epithelial ovarian cancer undergoing radical resection. J Int Med Res. 
2012;40(5):1822-9. 
19. Shakhar G, Blumenfeld B. Glucocorticoid involvement in suppression of NK 
activity following surgery in rats. J Neuroimmunol. 2003; 138(1-2):83-91.  
20. Votta-Velis E, Piegeler T, Minshall R, Aguirre J, Beck-Schimmer B, 
Schwartz D, et al. Regional anaesthesia and cancer metastases: The 
implication 
of 
local 
anaesthetics. 
Acta 
Anaesthesiol 
Scand. 
2014;57(10):1211-29 . 
21. Benzonana LL, Perry NJ, Watts HR, Yang B, Perry IA, Coombes C, et al. 
Isoflurane, a commonly used volatile anesthetic, enhances renal cancer 
growth and malignant potential via the hypoxia-inducible factor cellular 
signaling pathway in vitro. Anesthesiology. 2013; 119(3):593-605.  
22. Lennon FE, Mirzapoiazova T, Mambetsariev B, Salgia R, Moss J, 
Singleton PA. Overexpression of the μ-opioid receptor in human non-small 
cell lung cancer promotes Akt and mTOR activation, tumor growth, and 
metastasis. Anesthesiology. 2012; 116(4):857-67.  
23. Krog J, Hokland M, Ahlburg P, Parner E, Tønnesen E. Lipid solubility- 
and concentration-dependent attenuation of in vitro natural killer cell 
cytotoxicity by local anesthetics. Acta Anaesthesiol Scand. 2002; 
46(7):875-81.  
24. Ramirez MF, Tran P, Cata JP. The effect of clinically therapeutic plasma 
concentrations of lidocaine on natural killer cell cytotoxicity. Reg Anesth 
Pain Med. 2015;40(1):43-8.  

24. 
25. Reips UD, Funke F. Interval level measurement with visual analogue 
scales in Internet-based research: VAS Generator. Behav Res Methods. 
2008; 40:699–704. 
26. Spallone V, Morganti R, D’amato C. Validation of DN4 as a screening tool 
for neuropathic pain in painful diabetic polyneuropathy. Diabetic Med. 
2012; 29(5):578–85. 
27.    Boone H, Boone D. Analyzing likert data. J Ext. 2012;50(2):1–5. 
28.   Fields A, Huang J, Schroeder D, Sprung J, Weingarten T. Agitation in 
adults in the post-anaesthesia care unit after general anaesthesia. Br J 
Anaesth. 2018 Nov;121(5):1052-1058.