\documentclass[10pt]{article}

\usepackage{fullpage}
\usepackage{setspace}
\usepackage{parskip}
\usepackage{titlesec}
\usepackage[section]{placeins}
\usepackage{xcolor}
\usepackage{breakcites}
\usepackage{lineno}
\usepackage{hyphenat}





\PassOptionsToPackage{hyphens}{url}
\usepackage[colorlinks = true,
            linkcolor = blue,
            urlcolor  = blue,
            citecolor = blue,
            anchorcolor = blue]{hyperref}
\usepackage{etoolbox}
\makeatletter
\patchcmd\@combinedblfloats{\box\@outputbox}{\unvbox\@outputbox}{}{%
  \errmessage{\noexpand\@combinedblfloats could not be patched}%
}%
\makeatother


\usepackage{natbib}




\renewenvironment{abstract}
  {{\bfseries\noindent{\abstractname}\par\nobreak}\footnotesize}
  {\bigskip}

\titlespacing{\section}{0pt}{*3}{*1}
\titlespacing{\subsection}{0pt}{*2}{*0.5}
\titlespacing{\subsubsection}{0pt}{*1.5}{0pt}


\usepackage{authblk}


\usepackage{graphicx}
\usepackage[space]{grffile}
\usepackage{latexsym}
\usepackage{textcomp}
\usepackage{longtable}
\usepackage{tabulary}
\usepackage{booktabs,array,multirow}
\usepackage{amsfonts,amsmath,amssymb}
\providecommand\citet{\cite}
\providecommand\citep{\cite}
\providecommand\citealt{\cite}
% You can conditionalize code for latexml or normal latex using this.
\newif\iflatexml\latexmlfalse
\providecommand{\tightlist}{\setlength{\itemsep}{0pt}\setlength{\parskip}{0pt}}%

\AtBeginDocument{\DeclareGraphicsExtensions{.pdf,.PDF,.eps,.EPS,.png,.PNG,.tif,.TIF,.jpg,.JPG,.jpeg,.JPEG}}

\usepackage[utf8]{inputenc}
\usepackage[english]{babel}



\usepackage{float}








\begin{document}

\title{Factor Activity Levels and Bleeding Scores in Pediatric Hemophilia
Carriers Enrolled in the ATHNdataset.}



\author[1]{John Puetz}%
\author[2]{Brian Hu}%
\affil[1]{SSM Health Cardinal Glennon Children's Hospital}%
\affil[2]{American Thrombosis and Hemostasis Network}%


\vspace{-1em}



  
  \date{May 24, 2024}


\begingroup
\let\center\flushleft
\let\endcenter\endflushleft
\maketitle
\endgroup





\selectlanguage{english}
\begin{abstract}
\textbf{Background:} Multiple studies have now shown that a significant
proportion of hemophilia carriers meet criteria for having hemophilia
and/or report abnormal bleeding. However, to date, investigations of
hemophilia carriers have almost exclusively involved women over 18 years
of age. Little is known about factor activity levels and bleeding scores
in carriers during childhood. We queried a large deidentified database
of subjects with bleeding disorders residing in the United States to
determine factor activity levels and bleeding scores.
\textbf{Procedures:} The ATHNdataset was queried for hemophilia carriers
under 18 years of age. Collected information included demographics,
factor activity levels, bleeding scores. \textbf{Results:} Over 700
carriers in the pediatric age group were identified of which 626
submitted factor activity levels. Nearly half had factor activity levels
less than 40 IU/dL, thereby meeting criteria for having hemophilia. Of
those reporting bleeding scores, only 13.5\% reported an abnormal
bleeding score for age. The proportion reporting abnormal bleeding
scores was higher in those with factor levels less than 40 IU/dL (23\%)
than those greater than 40 IU/dL (9.7\%). \textbf{Conclusions:} The
proportion of pediatric carriers with hemophilia was double of that
previously reported for adults. Of those with hemophilia reporting a
bleeding score, the majority (77\%) did not report an abnormal bleeding
score for age. However, nearly 10\% of pediatric carriers not meeting
criteria for having hemophilia reported abnormal bleeding scores for
age. Similar results are reported in adults suggesting that factor
activity levels may not be predictive of bleeding symptoms in carriers.%
\end{abstract}%



\sloppy


\textbf{Introduction}

Hemophilia A and B are X-linked recessive conditions due to deficiencies
of factors VIII and IX respectively. Affected males are prone to
abnormal bleeding from trivial causes. Several studies over the past few
decades have shown that hemophilia carriers also have a bleeding
tendency.\textsuperscript{1-3} Due to random X chromosome inactivation,
female hemophilia carriers can have wide ranging factor activity
levels.\textsuperscript{2,4} Approximately one-fourth of hemophilia
carriers seen at hemophilia treatment centers (HTCs) have factor
activity levels less than 40 IU/dL and are now classified as having
hemophilia.\textsuperscript{1,4} Even amongst carriers seen at HTCs with
factor activity levels greater than 40 IU/dL, about one-fourth report
abnormal bleeding scores (BLS).\textsuperscript{1} Thus, amongst
hemophilia carriers, just less than one-half either have hemophilia
and/or report abnormal BLS.

To date, studies of hemophilia carriers have almost exclusively focused
on women over 18 years of age. Little is known about bleeding symptoms
of hemophilia carriers during childhood. Only one study has included a
significant sample size of children.\textsuperscript{1} It showed that
children were less likely than adults to have an abnormal BLS for age.
Additional details such as the proportion of children who had factor
levels less than 40 IU/dl, or the effects of race, ethnicity, type of
hemophilia, or mutation on BLS were not included.

The American Thrombosis and Hemostasis Network (ATHN) is an organization
whose purpose is to improve the lives of people with bleeding and
clotting disorders. To that end, ATHN sponsors a HIPPA-compliant,
de-identified dataset in which subjects agree to submit their health
information. Over 140 Hemophilia Treatment Centers (HTCs) across the
United States have collected and submitted data into the ATHNdataset.
Over 60,000 people with bleeding or clotting disorders have submitted
data. Of this, approximately 3300 are hemophilia carriers, including 743
in the pediatric age group.

The purpose of this investigation is to characterize hemophilia carriers
under 18 years of age enrolled in the ATHNdataset.

\textbf{Methods}

The ethical committees of participating HTCs authorized enrollment in
the ATHNdataset. The parents or guardians of eligible children either
opted in or provided informed consent to share their child's health
information. Core data elements, including demographics, primary
diagnosis, baseline factor activity levels, prescribed medications,
inhibitor status, and insurance are submitted by each
HTC.\textsuperscript{1} The International Society on Thrombosis and
Haemostasis Bleeding Assessment Tool Bleeding Score (ISTH-BAT BLS),
factor activity level, and genotype were collected for participants in
the \emph{My Life Our Future} (\emph{MLOF} ) genotype initiative.
Additional data elements such as bleeding events, including detailed
menstrual bleeding, medication usage, and joint range of motion could
also be submitted. Core data elements are audited by ATHN for
consistency. Many other data element's collection and submission are
left to the discretion of participating HTCs and are not audited.
Because data elements such as bleeding events, medication usage, or
detailed menstrual bleeding are likely underreported, this type of data
was not subjected to further analysis.

The ATHNdataset was queried in June 2022 for hemophilia carriers under
18 years of age. Collected data included age, race, ethnicity, type of
hemophilia (A or B), baseline factor activity level, genotype, and
ISTH-BAT BLS. The ISTH-BAT BLS has been validated for use in children,
and normal ranges are established for adults and
children.\textsuperscript{5} An ISTH-BAT BLS of 3 or higher was
determined to be abnormal for children under 18 years of age by
Elbatarny et al.\textsuperscript{5} Although Elbatarny et al. initially
established an ISTH-BAT score of 6 or higher as abnormal in adult women,
subsequent analysis has shown a ISTH-BAT score of 5 or higher was
abnormal for women aged 18-30.\textsuperscript{6} Since adolescents are
biologically more similar to women age 18-30 (menstruating and at risk
for post-partum hemorrhage) than children, we felt the revised
definition of an abnormal ISTH-BAT score was more appropriate to use in
adolescents aged 11-17. Doherty et al. did not revise the abnormal BLS
for adolescents due to sample size
limitations.\textsuperscript{6}Additional support for this methodology
is found in Jain et al.\textsuperscript{7} This study showed that a BLS
of 5 or higher in adolescent girls was predictive of having a bleeding
disorder. As previously described, we defined a BLS of 5 or higher as
abnormal for adolescents and 3 or higher as abnormal for
children.\textsuperscript{1}The participant's age at the time of BLS
determination was used to determine if the score was abnormal or not.
For subjects with multiple reported ISTH-BAT BLS we selected the
earliest record. The one-stage factor activity level is the predominant
methodology used to determine factor activity levels in the United
States, and factor activity levels reported in this study were presumed
to be from a one-stage assay. For subjects submitting more than one
factor activity level, the lowest reported level (baseline) was used.
The proportion of subjects who had a genotype determined was collected
to describe the population. A detailed analysis of genetic information
was beyond the scope of this study. Because this updated query covered a
different timeline than prior studies of hemophilia carriers using the
ATHNdataset, sample size differences were expected.

Descriptive statistics were used to define the population. Data sets
with a sample size less than 50 were considered to be too small for
valid statistical comparisons. Pearson's Chi squared test, Fisher's
exact test, Wilcoxon rank sum and Welch Two Sample t-tests were used for
group comparisons. When able to be measured, a p value of less than 0.05
was considered statistically significant. The original, de-identified
data can be obtained by contacting ATHN at support@athn.org.

\textbf{Results}

As of June 2022, 743 females under 18 years of age who were identified
as having hemophilia A or B or were carriers of hemophilia A or B were
enrolled in the ATHNdataset. Demographic information regarding these
subjects is found in table 1. P values for all comparisons were
\textgreater{}0.05. Three-hundred-forty-three were children under 11
years of age and 400 were adolescents between 11 and 17.99 years of age.
Roughly three-fourths were hemophilia A carriers or had hemophilia A,
and one-fourth were hemophilia B carriers or had hemophilia B. Of the
626 with known factor activity levels, 305 (49\%) had activity levels of
40 IU/dL or less, thereby meeting criteria for having hemophilia. Of
these, 18 had activity levels in the severe range, and 9 were moderate.
Adolescents with hemophilia B were more likely to report factor activity
levels less than 40 IU/dL than adolescents with hemophilia A (p=0.02).
There was no statistical difference in children reporting factor
activity levels \textless{} 40 IU/dL between hemophilia A and B (p=0.3).
The mean factor activity (standard deviation) for children was 45 (28)
and for adolescents was 47 (28) (p=0.5). Figure 1 shows the distribution
of factor activity levels for the entire pediatric population of
subjects enrolled in the ATHNdataset.

Two-hundred-twenty-three subjects reported an ISTH-BAT BLS and
demographics regarding this population is found in table 2. Ninety-eight
percent of subjects reporting a BLS participated in the
\emph{MLOF}genotype initiative. The mean BLS for adolescents (1.35)
compared to children (0.86), was statistically different (p=0.03), but
not a clinically relevant difference. Only 30 (13.5\%) subjects reported
an abnormal bleeding score for age. Seventeen percent of adolescents
reported an abnormal BLS for age compared to 7.7\% of children
(p=0.045). There was no statistically significant difference in those
reporting an abnormal BLS between hemophilia A and B for children or
adolescents (p=0.3 and p=0.8). Of the 223 subjects who submitted an
ISTH-BAT BLS, 195 submitted both a BLS and factor activity level.
Twenty-seven (13.8\%) reported an abnormal BLS for age. Figure 2 shows
the proportion with an abnormal bleeding score by factor activity level
for children and adolescents. Fourteen (23\%, 3 children) subjects with
factor activity levels under 40 IU/dL reported an abnormal BLS for age
while 13 (9.7\%, 4 children) of those with factor activity levels over
40 IU/dL did. Because the sample size of subjects reporting an abnormal
BLS was so small, additional evaluations to determine factors
contributing to an abnormal BLS were not undertaken.

\textbf{Discussion}

Previous investigations of adult hemophilia carriers, including an
investigation of the ATHNdataset, have shown that roughly one-fourth
meet criteria for having hemophilia.\textsuperscript{1,4} Our
investigation of pediatric carriers enrolled in the ATHNdataset showed
nearly half of pediatric carriers meet current criteria for having
hemophilia. There are several potential explanations for the high
proportion of pediatric carriers in this study having hemophilia. One is
that there could be ascertainment bias in the ATHNdataset. Pediatric
carriers with hemophilia (and more likely to have abnormal bleeding) may
be more likely to be seen at a HTC, thereby skewing the data. However,
of those reporting a BLS, the majority reported a normal BLS for age,
and the median BLS was 0. This suggests that recruitment of subjects
with a higher bleeding risk may not be skewing the data to a significant
extent. Also, similar ascertainment bias may exist in studies of adult
carriers. Pre-analytical or analytical errors are another possible
explanation. However, these specimens were handled by HTCs well versed
in handling blood specimens for hemophilia patients. Also, many of the
subjects in this study are seen at the same HTCs that also see adults.
Factor VIII is an acute phase response protein and if anything, the
stress of phlebotomy in a child might lead to falsely elevated factor
VIII activity levels. Pre-analytical and analytical issues are unlikely
to explain the high proportion of pediatric carriers with hemophilia
enrolled in the ATHNdataset compared to adult carriers with hemophilia
enrolled in the same database. It is known that factors VIII and IX
levels rise with age, and the mean factor activity level in child and
adolescent carriers was lower than that reported for
adults.\textsuperscript{2,8,9} This could explain the difference in
proportion of those reporting a factor activity below 40 IU/dL in adults
compared to pediatric carriers.

Of those reporting a BLS, a minority (9.7\%) of pediatric carries with
factor activity levels \textgreater{}40 IU/dL in the ATHNdataset report
BLS that were abnormal for age. Abnormal bleeding with normal factor
activity levels is a consistent finding in multiple studies of adult
carriers as well and seems to be more prevalent than in
children.\textsuperscript{1-3} Again, it is possible that bias could
explain this finding. A very small, single institution study showed a
higher BLS for carriers that knew their carrier status prior to
obtaining their BLS compared to carriers that did
not.\textsuperscript{10} In addition, cultural difference in the
reporting of BLS in carriers have been
reported.\textsuperscript{1,11,12} It is possible that carriers with
normal factor activity levels do not truly have abnormal bleeding, and
the apparent abnormal BLS are due to methodologic/cultural issues used
to determine the BLS. However, this should not be assumed to be the
explanation. To date, studies demonstrating a strong correlation between
factor VIII and IX activity levels and bleeding risk have been done
exclusively in males. It is not unreasonable to propose that factors
VIII and/or IX have gender related hemostatic functions that are not
measured by aPTT based assays. Factor VIII, at least, is known to have a
non-hemostatic function that is not measured by an aPTT based
assay.\textsuperscript{13} The factor VIII:Von Willebrand complex is
important for bone homeostasis, and female carriers have recently been
shown to have higher rates of osteoporosis and fractures compared to an
unaffected population.\textsuperscript{13,14} In factor XI deficiency,
there is a no correlation between factor activity levels and clinical
bleeding.\textsuperscript{15} It has been proposed that this is due to
hemostatic effects of factor XI that are not measured by aPTT based
assays.\textsuperscript{16,17} As shown in table 3, moderate to weak
correlation between factor activity and clinical bleeding is seen in
other coagulation factor deficiencies.\textsuperscript{15,18} Additional
investigation into possible hemostatic functions of factor VIII and IX
outside those measured by aPTT based assays are needed.

Another finding of our study was that most pediatric females with
hemophilia that reported a BLS had a normal BLS for age. This is also
seen in adult carriers with hemophilia.\textsuperscript{1} However, an
overwhelming majority with hemophilia had factor activity levels between
16 IU/dL and 40 IU/dL, levels that are expected to have a lower risk of
abnormal bleeding.\textsuperscript{19} Alternatively, this could be
another example of the inadequacy of aPTT based factor assays in
predicting bleeding symptoms in this population.

Several guidelines recommend against testing potential carriers for
genetic diseases during childhood.\textsuperscript{20.21} A principal
reason for this stems from ethical concerns regarding the loss of the
child's future autonomy. However, these guidelines have not met with
universal acceptance.\textsuperscript{22} A more recent policy statement
from British Medical Health allows for carrier testing so long as no
harm comes to the child.\textsuperscript{22} The American Academy of
Pediatrics and American College of Medical Genetics most recent
guideline also recommends against carrier testing unless a child is at
risk for childhood onset conditions.\textsuperscript{21} Our study
suggests that hemophilia carriers are at significant risk for having
hemophilia and/or abnormal bleeding. Because factor activity levels may
not be diagnostic of carrier status and may not be predictive of
bleeding risk, genetic testing of potential/obligate carriers seems
indicated.

This study has several strengths, even compared to other investigations
of adult carriers, including data submitted from multiple institutions,
genotyping of a significant proportion of subjects, standardized measure
of bleeding symptoms, and a sample size that is larger than most studies
involving adult hemophilia carriers.\textsuperscript{2,3,23} Even with
this large sample size, only a small number of pediatric carriers
reported an abnormal BLS for age. Thus, we could not make valid analysis
of other factors that might contribute to an abnormal BLS. Further
improvement of the study would have been inclusion of additional details
regarding clinical bleeding such as joint or menstrual bleeding.
However, this type of data is not consistently submitted by
participating HTCs and would lead to an underestimation of bleeding
symptoms.

Our investigation suffers from a weakness common to many previous
studies of hemophilia carriers, namely, they only investigate carriers
seen at a HTC. It is unknown to what degree carriers seen at a HTC are
representative of all hemophilia carriers, most of whom are not seen at
a HTC. The majority of subjects in this study did not submit a BLS, and
of those that did, nearly all were participants in the
\emph{MLOF}genotype initiative. Despite this limitation, the sample size
of pediatric subjects that did submit a BLS was larger than most studies
of adult carriers that report BLS, thereby making this data of value.

We relied on the ISTH-BAT BLS to describe bleeding symptoms. Bleeding
scores remain the best objective measure of clinical bleeding and have
been validated for carriers.\textsuperscript{23} However, there are
limitations, which may be particularly relevant for pediatric hemophilia
carriers. For instance, a 2-year-old carrier with a single untreated
joint bleed and no other abnormal bleeding would get an ISTH-BAT BLS
score of 2. This would not flag as abnormal for age in this study, but
most clinicians would consider this abnormal bleeding. Unlike Von
Willebrand disease, investigations of carriers have noted a poor
correlation between factor activity levels and bleeding
symptoms.\textsuperscript{1, 24-26} It is unknown if this is due to the
inadequacies of the BLS, factor activity levels, or both.

The benefits of a large sample size can be offset by errors associated
with large databases, including enrollment, data collection and
submission, and query errors. We attempted to limit this by restricting
this investigation to continuous variables (factor activity and BLS) and
categorical variables that are audited by ATHN. This study was limited
to pediatric carriers residing in the United States who participated in
the ATHNdataset and may not be reflective of pediatric carriers residing
in other areas of the world.

Despite the limitations of this study, as one of the few investigations
of hemophilia carriers focusing exclusively on the pediatric population,
we believe it makes important contributions. Significant knowledge gaps
remain regarding clinical bleeding in this population and additional
investigation is needed. Also needed are investigations into the
potential hemostatic functions not measured by aPTT based assays. This
would have applicability beyond hemophilia carriers and include all
patients with coagulation factor deficiencies.

\textbf{Acknowledgments} : The authors would like to acknowledge ATHN
and all the participating hemophilia treatment centers including
physicians, nurse practitioners, nurses, and data managers for
collecting and submitting data.

\textbf{Conflicts of Interest:} The authors report no conflicts of
interest for this project.

\textbf{References:}

\begin{enumerate}
\tightlist
\item
  Puetz J, Cheng D. Descriptive analysis of bleeding symptoms in
  haemophilia carriers enrolled in the ATHNdataset.
  \emph{Haemophilia.}2021;27(6):1045-1050.
\item
  Paroskie A, Gailani D, DeBaun MR, Sidonio RF Jr. A cross-sectional
  study of bleeding phenotype in haemophilia A carriers. \emph{Br J
  Haematol.} 2015;170(2):223-8.
\item
  Plug I, Mauser-Bunschoten EP, Brocker-Vriends AH et al. Bleeding in
  carriers of hemophilia. \emph{Blood} . 2006;108(1):52-6
\item
  Van Galen K, d'Oiron R, James P et al. A new hemophilia carrier
  nomenclature to define hemophilia in women and girls: Communication
  from the SSC of the ISTH. \emph{J Thromb Haemost.}2021;19(8):1883-188.
\item
  Elbatarny M, Mollah S, Grabell J et al. Normal range of bleeding
  scores for the ISTH-BAT: adult and pediatric data from the merging
  project. \emph{Haemophilia.} 2014;20(6):831-5.
\item
  Doherty D, Grabell J, Christopherson P et al. Variability in
  International Society on Thrombosis and Haemostasis=Scientific and
  Standardization Committee endorsed Bleeding Assessment Tool (ISTH-BAT)
  score with normal aging in healthy females:contributory factors and
  clinical significance. \emph{J Thromb Haemost} 2023:21:880-886.
\item
  Jain S, Zhang S, Acosta M, Malone K, Kouides P, Zia A. Prospective
  evaluation of ISTH-BAT as a predictor of bleeding disorder in
  adolescents presenting with heavy menstrual bleeding in a
  multidisciplinary hematology clinic. \emph{J Thromb
  Haemost}2020;18(10):2542-2550.
\item
  Biguzzi E, Castelli F, Lijfering W et al. Rise of levels of von
  Willebrand factor and factor VIII with age: Role of genetic and
  acquired risk factors. \emph{Thromb Res} 2021;197:172-178.
\item
  Appel I, Grimminck B,GeertsJ et al. Age dependency of coagulation
  parameters during childhood and puberty. \emph{J Thromb Haemost} 2012;
  10:2254-2263.
\item
  Puetz J, Johnson L, Hugge C, Buchanan P. First Look at My Life, Our
  Future Carrier Data from a Pilot Site. \emph{Res Pract Thromb
  Haemost.} 2018;2(S1):60.
\item
  Young JE, Grabell J, Tuttle A et al. Evaluation of the
  self-administered bleeding assessment tool (Self-BAT) in haemophilia
  carriers and correlations with quality of life.
  \emph{Haemophilia.}2017;23(6):e536-e8.
\item
  Li S, Fang Y, Li L et al. Bleeding assessment in haemophilia
  carriers-High rates of bleeding after surgical abortion and
  intrauterine device placement: A multicentre study in
  China.\emph{Haemophilia.} 2020;26(1):122-8.
\item
  Baud'huin M, Duplomb L, Teletchea S et al. Factor VIII-von Willebrand
  Factor Complex Inhibits Osteoclastogenesis and Controls Cell
  Survival.\emph{J Biol Chem} 2009;284(46)31704-31713.
\item
  Citla-Sridhar D, Sidonio R, Ahuja S. Bone health in haemophilia
  carriers and persons with von Willebrand disease: A large database
  analysis. \emph{Haemophilia} 2022;28(4):671-678.
\item
  Saes J, Verhagen M, Meijer K et al. Bleeding severity in patients with
  rare bleeding disorders: real-life data from the RBiN
  study.\emph{Blood Adv} 2020;4(20):5025-5034.
\item
  Puetz J, Hugge C, Moser K. Normal aPTT in children with factor XI
  deficiency. \emph{Pediatr Blood Cancer} 2018;65(4):e26910
\item
  Wheeler A, Gailani D. Why factor XI deficiency is a clinical
  concern.\emph{Expert Rev Hematol} 2016;9(7):629-637.
\item
  Peyvandi F, Palla R, Menegatti M et al. Coagulation factor activity
  and clinical bleeding severity in rare bleeding disorders: results
  from the European Network of Rare Bleeding Disorders. \emph{J Thromb
  Haemost} 2012;10(4):615-621.
\item
  Den Uijl I, Bunschoten E, Roosendaal G et al. Clinical severity of
  haemophilia A: does the classification of the 1950s still
  stand?\emph{Haemophia} 2011;17(6):849-853.
\item
  Borry P, Fryns J, Schotsmans P, Dierickx K. Carrier testing in minors:
  a systematic review of guidelines and position papers. \emph{Eur J Hum
  Genet} 2005;14(2):133-138.
\item
  Committee on Bioethics, Committee on Genetics, and The American
  College of Medical Genetics and Genomics Social, Ethical and Legal
  Issues Committee. Ethical and Policy Issues in Genetic Testing and
  Screening of Children.\emph{Pediatrics} 2013;131(3):620-622.
\item
  Vears D, Metcalfe S. Carrier testing in children and
  adolescents.\emph{Eur J Hum Genet} 2015;58(12):659-667.
\item
  James PD, Mahlangu J, Bidlingmaier C et al. Evaluation of the utility
  of the ISTH-BAT in haemophilia carriers: a multinational
  study.\emph{Haemophilia.} 2016;22(6):912-8.
\item
  Tosetto A, Rodeghiero F, Castaman G et al. A quantitative analysis of
  bleeding symptoms in type 1 von Willebrand disease: results from a
  multicenter European study (MCMDM-1 VWD). \emph{J Thromb
  Haemost.}2006;4(4):766-73.
\item
  Olsson A, Hellgren M, Berntorp E, Ljung R, Baghaei F. Clotting factor
  level is not a good predictor of bleeding in carriers of haemophilia A
  and B. \emph{Blood Coagul Fibrin.} 2014;25(5):471-5.
\item
  Miesbach W, Alesci S, Geisen C, Oldenburg J. Association between
  phenotype and genotype in carriers of haemophilia A.\emph{Haemophilia}
  2011;17(2):246-251.
\end{enumerate}

\textbf{Legends:}

Figure 1. Shows the distribution of factor activity levels for all
pediatric carriers enrolled in the ATHNdataset.

Figure 2. Shows the percentage with an abnormal BLS by factor activity
for child and adolescent hemophilia carriers.\selectlanguage{english}
\begin{figure}[H]
\begin{center}
\includegraphics[width=0.70\columnwidth]{figures/pbc-carr-fig-1/pbc-carr-fig-1}

\end{center}
\end{figure}\selectlanguage{english}
\begin{figure}[H]
\begin{center}
\includegraphics[width=0.70\columnwidth]{figures/pbc-carr-fig-2/pbc-carr-fig-2}

\end{center}
\end{figure}

\textbf{Hosted file}

\verb`pbc carr table 3.docx`  available at \url{https://authorea.com/users/616405/articles/642425-factor-activity-levels-and-bleeding-scores-in-pediatric-hemophilia-carriers-enrolled-in-the-athndataset}

\textbf{Hosted file}

\verb`subgroup table 1_updated_bh .docx`  available at \url{https://authorea.com/users/616405/articles/642425-factor-activity-levels-and-bleeding-scores-in-pediatric-hemophilia-carriers-enrolled-in-the-athndataset}

\textbf{Hosted file}

\verb`subgroup table 2_updated_bh.docx`  available at \url{https://authorea.com/users/616405/articles/642425-factor-activity-levels-and-bleeding-scores-in-pediatric-hemophilia-carriers-enrolled-in-the-athndataset}

\selectlanguage{english}
\FloatBarrier
\end{document}

