Clinical trial NCT05530252
Conditions: Periodontitis
Sponsor: Beijing Stomatological Hospital, Capital Medical University
Start date: 2022-09-05; Completion date: 2023-01-30
Enrollment: 51 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Chronic periodontitis is an infectious disease of periodontal support tissues caused by bacterial biofilm, which leads to inflammation and destruction of periodontal support tissues ultimately resulting in tooth loss.In the clinic, patients with Stage III and Grade B periodontitis are difficult to gain desired outcomes on account of deep periodontal pockets, complicated anatomy, the removal of subgingival dental biofilms, and control of residual inflammation. A large number of studies have indicated that the effectiveness of local application of antibiotics as an adjunct to scaling and root planning (SRP), such as the antimicrobial and minocycline hydrochloride could affect bacterial metabolism and inhibit bioﬁlm attachment particularly in terms of pocket depth reduction and attachment level gain. However, the use of wide-spectrum antibiotics may cause some inevitable side effects including drug resistance, pathogens and probiotics were eliminated leading to diversity of microbiota diminished, and toothstaining. To solve the problems of antibiotics in the clinic, antimicrobial peptides (AMPs) may be considered as an alternative to conventional antibiotics drugs.In this randomized clinical trial, we aimed to evaluate the effects of AMPs as an addition to SRP on clinical parameters and microbiological biofilms in patients with Stage III and Grade B periodontitis.

Detailed description:

Arms: arm 1: scaling and root planning and subgingival application of antimicrobial peptide gel arm 2: scaling and root planning and subgingival application of minocycline hydrochloride ointment arm 3: scaling and root planning
Interventions: intervention 1: Oral biological antimicrobial gel intervention 2: minocycline hydrochloride ointment intervention 3: SRP
Intervention descriptions: intervention 1: The patients were divided into subgingival scaling, and Oral Biological Antimicrobial Gel was used in the periodontal pocket after scaling. The injection probe with gel was inserted into the periodontal pocket, and the injection was slowly pushed, and the injection was gradually withdrawn during the injection, until a little gel was found at the mouth of the bag. intervention 2: The patients were divided into subgingival scaling, and minocycline hydrochloride ointment was used in the periodontal pocket after scaling. The injection probe with gel was inserted into the periodontal pocket, and the injection was slowly pushed, and the injection was gradually withdrawn during the injection, until a little gel was found at the mouth of the bag. intervention 3: only scaling and root planning was performed on the patient

Protocol documents:
Prot_SAP_000.pdf: 
Effects of Antimicrobial Peptides Application after Non-surgical 
Periodontal Therapy on Treatment of Stage III and Grade B 
Periodontitis 
 
The Medical Ethics Committee of Beijing Stomatological Hospital 
Capital Medical University approved the clinical study protocol 
(CMUSH-IRB-KJ-PJ-2018-03) 
Clini calTr ials. gov Identifier:NCT05530252 
Date:2022.09 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


Contents 
Part one:  
The research purpose and project basis  
I. 
Project basis  
II. 
Purpose of research  
Part two:  
Clinical trial protocol  
I. 
Objects and source   
II. 
Estimate of sample size  
III. 
Measures  
IV. 
Clinical trial procedure  
Part three:  
Research object rights and adverse event treatment  
I. 
Research object rights and adverse event treatment  
II. 
Informed consent process  
Part four:  
Statistical analysis plan  
  
  
 
 
 


Part one:  
The research purpose and project basis  
1.Project basis  
     Chronic periodontitis is an infectious disease of periodontal support tissues caused 
by bacterial biofilm, which leads to inflammation and destruction of periodontal support 
tissues ultimately resulting in tooth loss. In 2018, EEP and AAP sorted out periodontitis 
into four classifications (Stage I to IV) based on several variables including clinical 
attachment loss, amount and percentage of bone loss, probing depth, presence and 
extent of angular bony defects and furcation involvement, tooth mobility, and tooth loss 
due to periodontitis, and three levels (grade A: low risk, grade B: moderate risk, grade 
C: high risk of progression) according to the rate of disease progression. In the clinic, 
patients with Stage III and Grade B periodontitis are difficult to gain desired outcomes 
on account of deep periodontal pockets, complicated anatomy, the removal of 
subgingival dental biofilms, and control of residual inflammation. A large number of 
studies have indicated that the effectiveness of local application of antibiotics as an 
adjunct to scaling and root planning (SRP), such as the antimicrobial and minocycline 
hydrochloride could affect bacterial metabolism and inhibit biofilm attachment 
particularly in terms of pocket depth reduction and attachment level gain. However, the 
use of wide-spectrum antibiotics may cause some inevitable side effects including drug 
resistance, pathogens and probiotics were eliminated leading to diversity of microbiota 
diminished, and toothstaining. To solve the problems of antibiotics in the clinic, 
antimicrobial peptides (AMPs) may be considered as an alternative to conventional 
antibiotics drugs. Many studies reported that AMPs have distinct advantages in specific 
selectivity for bacterial targets and minimal emergence of microbial resistance in 
comparison with antimicrobial agents. Gingival epithelial as a biological barrier has the 
characteristic of specificity in the natural immune system and could produce AMPs 
including the expression of β-defensins, hBD-1, hBD-2and hBD311, and a large number 
of α-defense and LL-37 exist in the neutrophils of gingival sulcus, which play an 


important role in protecting gingival epithelial. However, the efficacy of antimicrobial 
peptide in the treatment of Stage III and Grade B periodontitis lack of more clinical 
studies.  
2.Purpose of research  
  In this randomized clinical trial, we aimed to evaluate the effects of AMPs as an 
addition to SRP on clinical parameters and microbiological biofilms in patients with  
Stage III and Grade B periodontitis.  
Part two:  
Clinical trial protocol  
1. Objects and source  
   Subjects were recruited from patients with Stage III and Grade B periodontitis from 
the periodontal department of Beijing Stomatological Hospital from Sep. 2022 to Oct.  
2022.  
2. Estimate of sample size  
Multiple test indexes were involved in this experiment, and the main test index was the 
decrease value of the average probing depth, which was intended to be referred to. 
Existing literature studies have shown that the local application of minocycline 
hydrochloride ointment can reduce the average periodontal pocket depth by 1.5mm, and 
it is expected that biological antibacterial peptides can reduce the average periodontal 
pocket depth by about 2mm.Set α=0.05, test efficiency, that is, 1-β was 90%, and the 
ratio of two samples was 1:1. According to the statistical formula of sample size  
estimation of two independent samples: n1=n2=2
, the sample size of each 
group could be calculated as 14 cases. Assuming that the shedding rate was 20%, the 
required sample size of each group was 17 cases, and the total sample number was 51 
cases.  
3.Measures  


Periodontal probing depth (PD) mm: from the gingival margin to the bottom of the 
periodontal pocket.  
Attachment level (AL)（mm）: from the cementoenamel junction, crown margin or 
restoration to the bottom of the pocket.  
Bleeding index (BI): the rating method was assessed using the criteria of Mazza (Mazza, 
Newman, & Sims, 1981).   
Microbiological sampling:(1) Sample site selecting: Four samples of subgingival plaque 
were collected separately from each subject: from the sites which following conditions: 
5 mm≦ PD≦ 10 mm, PI < 1, Mov≦II°, samples were taken at the same sites at 7 and 
90 days. (2) Plaque sampling: Dried samples teeth, remove the supragingival plaque 
with a probe, gently put the sterile Mini Grace scaler into the periodontal pocket, and 
take the subgingival plaque. The plaque sample was placed in a 1.5ml sterile centrifuge 
tube containing 1ml TE buffer (10mM Tris-HCl, 1mMEDTA; pH 8). (3) Samples 
storing: Specimens were collected and placed on ice immediately after sampling, 
transported to the laboratory within 15min and stored at −80°C.  
Microbial DNA sequencing  
Samples were conveyed on dry ice to the laboratory at Beijing Allwegene Tech, Ltd  
(Beijing, China) within 5 h. The microbial genomic DNA was isolated, amplified by 
PCR, and then the V3–V4 hypervariable regions of the 16S rRNA gene18 were subjected 
to high-throughput sequencing using the Illumina Miseq PE300 sequencing platform 
(Illumina, Inc. CA, USA).  
  
  
  
  
  
  


  
4.Clinical trial procedure  
 
 


Part three:  
Research object rights and adverse event treatment 
1.Research object rights and adverse event treatment object 
rights:  
a. All examinations and sample collection in this experiment were conducted by 
medical standard, and there was no trauma to the oral tissues and the whole body of the 
subjects.  
b. All operations in this study were routine operations for the treatment of stage III 
grade B periodontitis, and there was no experimental treatment.  
c. Minocycline hydrochloride and antibacterial polypeptide gel used in this research 
institute have been widely used in the treatment of periodontitis in our hospital to ensure 
their safety, and the costs of the two preparations are borne by the subject.  
d. The subject's participation in the study is voluntary, and the subject may refuse to 
participate in the study or withdraw from the study at any time without any penalty or 
loss of his or her due interest.  
e. To the extent permitted by applicable laws and/or regulations, records relating to 
the identification of subjects shall be kept confidential and shall not be made public. In 
the event of public publication of test results, the identity of subjects shall remain 
confidential  
f. 
Since this study is also a regular procedure for the treatment of Stage III Grade B 
periodontitis, treatment for this disease can continue if any subjects withdraw from the 
study.  
g. If there is any injury related to the test, the subject will be treated by a professional 
doctor and other alternative treatment options will be selected. The following are the 
possible injury events and treatment methods of the test.  
h. Subject or his/her legal agent will be notified in a timely manner if information 
becomes available that may affect the subject's continued participation in the study.  
adverse event treatment:  


This treatment belongs to the routine treatment of stage III Grade B periodontitis. 
Possible adverse reactions during and after treatment and emergency treatment by 
doctors are as follows:  
a. Anesthesia complications: subgingival curetting is a painless treatment, which 
requires local injection of anesthesia drugs before treatment. Possible complications of 
anesthesia include syncope, allergy, poisoning and hematoma. Emergency treatment 
methods are as follows:  
Syncope: Prevention of syncope: Reducing the psychological and physical stress of the 
patient is essential to prevent syncope. Do a good job of ideological work before 
treatment, eliminate tension, avoid operating on an empty stomach  
Management of syncope: 1) Stop medical procedures and remove foreign bodies in the 
mouth.2) Accurate monitoring of vital signs, focusing on monitoring patients' pulse 
(intensity and rhythm) and blood pressure: when the heart rate is lower than 60 beats 
per minute, the pulse is weak; When blood pressure is below 6.67 kPa (50 mmHg), it is 
generally difficult to reach a pulse.3) Patients with hypotension should change their 
body position to supine position and elevate their lower limbs; Loosen collar, oxygen; 
Ammonia stimulation increases ventilation.4) If symptoms do not improve, intravenous 
channels should be established quickly and Ringer's solution or normal saline should 
be injected at 250~500 mL per hour. If necessary, dopamine can be given to improve 
hypotension and atropine can improve bradycardia.5) Loss of consciousness is easy to 
lead to post lingual fall. When respiratory tract obstruction occurs, the lingual body and 
mandible should be pulled forward in time to open the respiratory tract.6) In case of 
cardiac or respiratory arrest, immediately perform cardiopulmonary resuscitation. 
Allergy: prevention of allergy: before anesthesia operation, ask patients in detail 
whether there is a history of local anesthetics allergic reaction, is the key to prevent 
allergic reaction, to ester local anesthetics allergic reaction and constitution of patients, 
are converted to amide drugs. If a patient has an adverse reaction to local anesthetics, it 
is important to consider whether the reaction is an allergic reaction and choose other 
local anesthetics or terminate the treatment.  


Allergy treatment: For mild allergic reactions, desensitization drugs such as calcium, 
promethazine, cortisone hormone intramuscular injection or intravenous injection and 
oxygen inhalation can be given; If allergic reaction is suspected, start treatment as soon 
as possible. The general principles of treatment are as follows.1) Change the body 
position so that the patient is supine and the lower limbs are elevated.2) Maintain airway 
patency and oxygen intake. When the patient has difficulty breathing, open the 
respiratory tract and give oxygen to the mask; If laryngeal edema occurs, endotracheal 
intubation or tracheotomy should be performed. In case of emergency, cricothyroi 
membrane puncture can be performed first; Bronchial spasm can be used to relieve 
spasm.3) Rapid establishment of intravenous channels and correct administration. 
Poisoning: 1) When central nervous system excitation reaction occurs, all operations 
should be stopped immediately, oxygen inhalation should be given, venous access 
should be quickly established, and the patient should be comforted.2) When patients 
have loss of consciousness, strict airway management should be conducted to keep the 
airway unobstructed.3) When patients have convulsions, oxygen should be given to 
prevent respiratory obstruction caused by vomiting; Anticonvulsants such as diazepam 
may be given intravenously if ventilation is affected.4) When circulatory system failure 
occurs, lower limbs should be elevated, venous perfusion should be rapid, and 
vasopressor drugs should be given  
Cardiac drugs such as dopamine, epinephrine, and isoproterenol.5) In case of 
respiratory and cardiac arrest, perform CPR immediately.  
Hematoma: If local hematoma appears, immediately stop injection, pressure to stop 
bleeding, give ice. In order to avoid local infection and hematoma enlargement, 
antibiotics and hemostatic drugs are given as appropriate. Avoid taking aspirin orally. 
After 48 hours local hot compress or physiotherapy, promote the absorption and 
dissipation of hematoma.  
b. Common adverse reactions after subgingival curetting include pain and bleeding. 
The pain and bleeding can be relieved spontaneously within 24H after treatment. If the 


pain reaction is large, accompanied by systemic inflammatory reaction, systemic 
antiinflammatory drugs should be taken. The trial was terminated.  
c. If other diseases in need of antibiotics occur during the test, in order to ensure the 
health of the test subjects, the test subjects can take antibiotics and terminate the test.  
d. Allergy to minocycline hydrochloride or antibacterial polypeptide gel used in this 
study: To ensure the maximum safety of subjects, subjects should be observed in the 
hospital for half an hour after treatment. In case of suspected allergic symptoms, 
immediately rinse the periodontal pocket with normal saline until all the preparations 
are flushed out. In subjects who developed allergies, the study was terminated and 
mechanical therapy alone was selected to complete the treatment of stage III grade B 
periodontitis.  
Procedures for withdrawing from the study  
The trial is terminated when:  
a. Subjects found not meeting the inclusion criteria after inclusion  
b. In case of serious adverse events or diseases requiring antibiotic administration 
during the study, the patient may voluntarily terminate the study in order to ensure 
the health of the study participant  
c. Subjects with poor compliance or inability to comply with test requirements  
d. It is not appropriate to continue the test for medical reasons or subjects' interests  
e. The subject asked to quit  
f. The patient is pregnant  
g. Patients who were combined with other drugs of the same type or used other 
treatments that affected the study during the trial  
h. Participate in other clinical investigators during the trial  
i. The researchers decided that the trial needed to be discontinued  
Case definitions for withdrawal from the study:  
All patients who fill in the informed consent form and are eligible to enter the trial have 
the right to withdraw from the clinical trial at any time, no matter when and for any 


reason, as long as the subject does not complete the observation period specified in the 
protocol, referred to as shedding cases.  
Treatment of shedding cases: After shedding patients, researchers should try their best 
to contact the patients, complete the assessment items that can be completed, and fill in 
the test conclusion form. For shedding due to adverse reactions, those who are 
determined by follow-up to be related to the treatment used in the study must be 
recorded in the case report form.  
Records of shedding cases: For any shedding case, the investigator must fill in the 
pathology report with reasons for shedding. Generally, there are 6 reasons for shedding: 
adverse events, lack of efficacy, loss of follow-up, subject withdrawal, investigator 
discontinuation, and others.  
2.Informed consent process  
a. Before agreeing to participate in the study, subjects should carefully read the 
informed consent, which has been approved by the Ethics Committee.If the informed 
consent is renewed, the subject or his/her legal representative will receive a new, 
signed and dated update approved by the Ethics Committee. This consent form is in 
duplicate, with one copy for the investigator and one copy for the subject.  
b. The informed consent process was carried out by Xie Yongmei, the researcher of this 
study, now working in the General Stomatology Department of Wangfujing 
Department of Beijing Stomatological Hospital.  
c. The explanation process of informed consent shall be narrated by the researcher in 
plain language that the subjects can understand. Vulnerable groups and groups 
without full capacity for civil conduct are not recruited in this experiment, including 
children, pregnant women, people with mental illness and cognitive disabilities, 
terminally ill patients and prisoners.  
d. In the process of informed consent, the researcher should give a detailed and clear 
explanation of the content that the subject does not understand or any question related 
to the experiment.  
e. Subjects must be given sufficient time to consider whether to participate in the study.  


f. Informed consent should be conducted in a quiet and separate environment to avoid 
pressure on the subject.  
g. Neither the investigator nor the relevant personnel of the study can coerce or 
improperly influence the subjects to make the decision of whether to participate in 
the study.  
h. The informed consent form shall be signed and dated by both the subject and the 
investigator performing the informed consent process.  
i. In the process of informed consent, researchers need to explain the following issues 
to subjects:  
(1) Purpose, duration, methods, costs, rights and inconvenience of subjects, expected 
benefits and risks of the study.  
(2) Inform subjects of the different groups to which they may be assigned and the 
control group of the potential benefits and risks.  
(3) Other current methods of diagnosis and treatment of this disease, and the possible 
benefits and risks of each method: Minocycline hydrochloride ointment and  
antibacterial polypeptide gel used in this treatment are sensitized. If the treatment plan 
of this test cannot be adopted, simple periodontal mechanical therapy can be chosen.  
(4) Inform the subject that participation in the study is voluntary, that the subject may 
refuse to participate or propose the study at any stage of the study without 
discrimination or retaliation, and that his medical treatment and rights will not be 
affected.  
(5) During the test, subjects will have access to relevant information at any time.  
(6) The Ethics Committee and relevant regulatory authorities shall be granted direct 
access to the subject's original medical records for the purpose of verifying the 
procedures and/or data of the clinical trial, to the extent permitted by applicable laws 
and regulations and without violating the subject's privacy.  
Part four:  
Statistical analysis plan  


All clinical indicator data were analyzed using IBM SPSS ver. 22.0, The data of normal 
distribution are represented by (𝑥̅±𝑠), while the data of non-normal distribution are 
represented by M (Q25, Q75). For the subjects' gender and age using chi-square analysis. 
First, the changes of PD, AL, BI at baseline and the three indicators after treatment were 
statistically analyzed, Secondly, PD reduction of anterior teeth, premolars, and molars 
after treatment were calculated. Finally, the reduced values of sites with different PD 
depths after treatment were analyzed. The above data satisfying the normal distribution 
were analyzed by ANOVA, and LSD was selected for comparison between two groups. 
Kruskal -- Wallis H test was adopted for the comparison between groups with 
nonnormal distribution. Tests with p < 0.05 were considered significant.  
Microbial sequencing data were processed using QIIME19 and MOTHUR20 by 
Allwegene Tech, Ltd. (Beijing, China). To retain only high-quality sequences for the 
downstream analysis, sequences that were less than a 100bpin length after quality 
trimming, contained one or more ambiguous base-calls (N), or had < 90% quality 
scores > Q20 were eliminated. After trimming, high-quality sequences were aligned to 
the Ribosomal Database Project and were clustered into operational taxonomical units 
(OTU) using QIIME at 97%similarity levels. Before further analysis, singleton OTUs 
were removed.  
  

ICF_001.pdf: 
Effects of Antimicrobial Peptides Application after Non-
surgical Periodontal Therapy on Treatment of Stage III and 
Grade B Periodontitis 
The Medical Ethics Committee of Beijing Stomatological 
HospitalCapital Medical University approved the clinical 
study protocol(CMUSH-IRB-KJ-PJ-2018-03) 
Clini calTr ials. gov Identifier:NCT05530252 
Date:2022.09 
Informed Consent Form 
You are invited to participate in this study, and this informed consent form can help 
you decide whether to participate in this study. Before you agree to join this study, 
please read the following carefully and ask the researcher carefully if you do 
not understand anything or have questions. 
1、 
Purpose and Content 
To evaluate the effects of the antimicrobial peptides (AMPs) as an adjunct to 
scaling and root planning (SRP) on clinical and microbiological outcomes in 
patients with periodontitis in Stage III and Grade B. 
2、 
Principal investigator: Yongmei Xie 
3、 
Research process 
The expected number of participants in the study is 51 for one year. You will be 
randomly assigned to either minocycline hydrochloride or biologic antibacterial 
peptides. The group you will be included in will not be of your choice or assigned by 
your physician, but will be determined by random methods. The periodontal 
examination, subgingival plaque sample collection and periodontal cleaning and 
curettage will be performed by the professional periodontist of our hospital, and 
minocyclic hydrochloride or biological antibacterial polypeptide preparation will be 
applied locally. Both periodontal examination and sample collection are non-invasive 
operations. Periodontal curettage is the basic treatment for periodontitis, and it is a 


painless debridement treatment when performed under local anesthesia. Individual 
studies take three months. 
Pre-experimental: A clinical-radiographic examination was carried out to determine the 
periodontal status of the subjects. Fifty-one patients with periodontitis met the inclusion 
criteria volunteered to participate after they were detailed about the purpose, adverse 
reactions, and efficacy of the study. The patient participants were randomly assigned 
into the treatment group, Perio group, and SRP group, then supragingival scaling and 
oral hygiene education were performed to them (Fig.1). 
Treatment: Seven days after the pre-experiment, the subjects were re-examined for 
periodontal clinical indicators recording and microbiological samples. After 
examination, subjects were assigned to receive one of the following treatments 
according to their groups: SRP group: only scaling and root planning, Perio group: 
scaling and root planning and subgingival application of minocycline hydrochloride 
ointment, or AMP group: scaling and root planning and subgingival application of 
antimicrobial peptide gel. The treatment was conducted by one experienced periodontist 
throughout the study. 
Re-examination: At 7 and 90 days, microbiological samples were collected repeatly at 
the selected sites, and clinical examination was performed 90 days after treatment. 
4、 Follow up 
The study was followed up twice after the subgingival curettage, one week after 
the end of the subgingival curettage and the second time three months after the end of 
the subgingival curettage. Follow-up included: sample collection, periodontal 
examination and oral education. The total duration of your participation in this study is 
three months. 


5、 
Safety and adverse reactions 
a. The drugs used in this study are all approved in China for the adjuvant 
treatment of clinical chronic periodontitis, purchased through the formal 
process of the hospital, with uniform concentration and purity, and widely 
used in the treatment of chronic periodontitis in the periodontal department of 
our hospital. 
b. Periodontal examination and treatment in this study were all conducted 
by attending doctors with rich undergraduate clinical experience. 
6、 
Subject's rights and interests 
Benefit: 
a. Professional periodontist will make personalized periodontal treatment 
plan and periodontal treatment for you. 
b. Get professional periodontal health education and maintenance. 
If you decide to participate in the study, you should: 
a. Follow up as planned, and do not stop scheduling appointments for treatment 
without first communicating with the study physician. 
b. Antibacterial drugs and antibiotics should not be used locally in the oral cavity 
during the study period, and any other drugs used during the study period should be 
reported to your doctor, including over-the-counter drugs purchased by you. c. Tell your 
doctor if you have any health problems. 
d. During the study period, oral hygiene should be maintained according to the oral 
hygiene maintenance methods taught by doctors 
e. If you fail to comply with the terms listed above, you may not participate in the 
study. 
Risk: 
a. Periodontal examination and sample collection are non-invasive and have no 
adverse effects on your oral tissue and general health. All the drugs used have been 
approved for the treatment of clinical chronic periodontitis to ensure safety. 
b. Periodontal treatment requires local anesthesia, which may result in the following 
complications: syncope, allergy, poisoning and hematoma. Please tell your doctor if you 


have a history of drug allergy or general illness. If there are special circumstances, the 
doctor will terminate the treatment and the trial. 
c. Common adverse reactions after subgingival curetting include pain and bleeding. 
The pain and bleeding can be relieved spontaneously within 24H after treatment. If the 
pain reaction is large, accompanied by systemic inflammatory reaction, systemic anti-
inflammatory drugs should be taken. The trial was terminated. 
Exit in the middle: 
a. You may withdraw from the study at any time. If you cannot comply with the study 
plan, or if the doctor considers it for medical or other reasons, he or she may decide to 
withdraw you. If you withdraw from the study for any reason, the doctor will ask you 
to return to complete the final procedures of the study b. The research process may be 
terminated if: 
Can not follow the time prescribed by the doctor 
Oral topical application of antibacterial mouthwash or drugs during the study period 
Confidentiality: 
All records showing your name will be kept confidential. Your name will never appear 
in published reports. 
Volunteer statement 
a. I voluntarily agreed to participate in this study. 
b. I have read and understood the risks set forth and described in this informed Consent. 
c. I knew I could withdraw from the study at any time. 
d. I have been given the opportunity to ask questions and I have understood the answers 
to all of them. 
Subject's signature: 
Investigator's signature: 
Date: