Preliminary investigation of human lymphatic malformations in vitro.
OBJECTIVE
To develop an in vitro model of human lymphatic malformations (LM) that reflects histological characteristics of native LM.


METHODS
Fresh human LM (n = 6) were harvested, sectioned, explanted into a fibrinogen gel, and cultured. A total of 25 explants were developed and observed for primary and peripheral cellular growth. On days 9 to 10, the cultured tissues and gels were collected and fixed in 10% formalin. Primary LM and surrounding gel matrix were sectioned and stained for H&E analysis. Immunohistochemistry was performed for Prox-1 and D2-40, known markers for lymphatic endothelium, and Ki-67, a marker of cellular proliferation.


RESULTS
On culture day 3, cells were observed to grow into the gel surrounding the primary tissue explants. Persistent and significant growth into the gel matrix was observed for each specimen at subsequent measurement intervals (day 6 and day 10, P < .0001). H&E staining of all the LM explants demonstrated survival and intact organization and cellular structure reflective of the original LM specimens. Microchannels were observed in the surrounding gel suggesting the presence of newly formed lymphatic vessels. Positive-immunohistochemical staining for D2-40 and Prox-1 revealed organized lymphatic endothelia within each specimen and associated microchannels distal to the explants in the gel matrix. Scattered cells in the gel matrix stained positive for Ki-67.


CONCLUSIONS
This experimental model suggests that human LM can be preserved and observed to grow in vitro with structural characteristics, and immunohistologic qualities similar to native LM. This model may provide a facile tool for basic and translational research on LM.
Department of Otolaryngology-Head and Neck Surgery, Division of Pediatric Otolaryngology and Center for the Investigation of Congenital Aberrancies of Vascular Development, University of Arkansas for Medical Sciences, Arkansas Children's Hospital, Little Rock, Arkansas 72207, USA.
Yuemeng
Y
Dai
Fang
F
Hou
Ali
A
Saad
Chun-Yang
CY
Fan
Lisa M
LM
Buckmiller
James Y
JY
Suen
Gresham T
GT
Richter
D000328:Adult; D049109:Cell Proliferation; D002648:Child; D002675:Child, Preschool; D004729:Endothelium, Lymphatic / Q000473:pathology*; D005260:Female; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D007124:Immunoenzyme Techniques; D007223:Infant; D019394:Ki-67 Antigen / Q000378:metabolism*; D044148:Lymphatic Abnormalities / Q000473:pathology* / Q000601:surgery; D008210:Lymphocele / Q000151:congenital / Q000473:pathology / Q000601:surgery; D008297:Male; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D046509:Tissue Culture Techniques; D014022:Tissue Survival / Q000502:physiology; D025521:Tumor Suppressor Proteins / Q000378:metabolism*; D000098551:Prospero-Related Homeobox 1 Protein
D018398:Homeodomain Proteins; D019394:Ki-67 Antigen; C472643:Knl1 protein, human; D008869:Microtubule-Associated Proteins; D025521:Tumor Suppressor Proteins; D000098551:Prospero-Related Homeobox 1 Protein