Bionic aggregation-induced emission photosensitizer for enhanced cancer immunotherapy.

Cold exposure therapy (CE), as an inexpensive method, has shown great potential in cancer therapy. Exploring the combined anti-tumor mechanism of CE and traditional therapies (such as photodynamic therapy (PDT)) is exciting and promising. Here, a bionic aggregation-induced emission photosensitizer system (named THL) is designed for combined CE to enhance anti-tumor immunotherapy. THL inherits the homologous targeting ability of tumor derived exosomes, promoting the enrichment of THL at the tumor site. Under external illumination, THL generates hydroxyl radicals and superoxide anions through type I PDT. In addition, mice are pretreated with cold exposure, which promotes THL mediated PDT and reactive oxygen species (ROS) generation by reducing the production of ATP and GSH in tumor tissue. This combination therapy increases production of ROS within the tumor, inhibits the growth of distant tumors, recurrent and rechallenged tumors and increases the number of cytotoxic CD8+T cells and memory T cells. Compared to PDT alone, combination therapy shows greater advantages in tumor immunotherapy. The combination therapy strategy provides new ideas for cancer immunotherapy.

Rocaglamide regulates iron homeostasis by suppressing hepcidin expression.

The anemia of inflammation (AI) is characterized by the presence of inflammation and abnormal elevation of hepcidin. Accumulating evidence has proved that Rocaglamide (RocA) was involved in inflammation regulation. Nevertheless, the role of RocA in AI, especially in iron metabolism, has not been investigated, and its underlying mechanism remains elusive. Here, we demonstrated that RocA dramatically suppressed the elevation of hepcidin and ferritin in LPS-treated mice cell line RAW264.7 and peritoneal macrophages. In vivo study showed that RocA can restrain the depletion of serum iron (SI) and transferrin (Tf) saturation caused by LPS. Further investigation showed that RocA suppressed the upregulation of hepcidin mRNA and downregulation of Fpn1 protein expression in the spleen and liver of LPS-treated mice. Mechanistically, this effect was attributed to RocA's ability to inhibit the IL-6/STAT3 pathway, resulting in the suppression of hepcidin mRNA and subsequent increase in Fpn1 and TfR1 expression in LPS-treated macrophages. Moreover, RocA inhibited the elevation of the cellular labile iron pool (LIP) and reactive oxygen species (ROS) induced by LPS in RAW264.7 cells. These findings reveal a pivotal mechanism underlying the roles of RocA in modulating iron homeostasis and also provide a candidate natural product on alleviating AI.

Euphohelioscopin A enhances NK cell antitumor immunity through GSDME-triggered pyroptosis.

Immune evasion by cancer cells poses a significant challenge for natural killer cell-based immunotherapy. Pyroptosis, a newly discovered form of programmed cell death, has shown great potential for enhancing the antitumor immunity of natural killer cells. Consequently, targeting pyroptosis has become an attractive strategy for boosting natural killer cell activity against cancer. In this study, various assays were conducted, including natural killer cell cytotoxicity assays, flow cytometry, xenograft tumor models, real-time polymerase chain reaction, and enzyme-linked immunosorbent assay, to assess natural killer cell-mediated cell killing, as well as gene and protein expressions. The results indicated that euphohelioscopin A, a potential pyroptosis activator, enhances natural killer cell-mediated lysis of tumor cells, resulting in inhibiting tumor growth that could be reversed by natural killer cell depletion. Furthermore, we found that euphohelioscopin A significantly enhanced IFNγ production in natural killer cells and synergistically upregulated GSDME with IFNγ in cancer cells. Euphohelioscopin A also increased the cleavage of GSDME, promoting granzyme B-induced pyroptosis, which could be reversed by GSDME knockdown and IFNγ blockade. Overall, the findings suggested that euphohelioscopin A enhanced natural killer cell-mediated killing of cancer cells by triggering pyroptosis, making euphohelioscopin A a promising pyroptosis activator with great potential for use in natural killer cell-based cancer immunotherapy.

[Low cement distribution index is a risk factor for refracture of the adjacent segments after percutaneous vertebroplasty].

OBJECTIVE: To investigate the impact of cement distribution index on the occurrence of refracture in the adjacent segments after percutaneous vertebroplasty. METHODS: This retrospective analysis was conducted among 143 patients who received percutaneous vertebroplasty for osteoporotic vertebral compression fracture between April, 2011 and April, 2014. Of the 134 patients with complete follow-up data, 18 had adjacent segment fracture within 1 year following the surgeries (re-fracture group), and 116 patients without new fracture served as the control group. All the patients underwent X-ray examinations after the surgery and according to the position and shape, the cement in the vertebrae were classified into 5 types (I to V), and the volume-cubage index was computed based on the cement volume and vertebral cubage. Age, gender, bone mineral density (BMD), cement distribution index, volume-cubage index, and cement leakage were evaluated in the 2 groups, and the variables with significant differences between the 2 groups were analyzed in Logistic regression analysis. RESULTS: BMD was significantly lower and the rate of cement leakage was significantly higher in the re-fracture group than in the control group (P<0.05). Significant difference was found in cement distribution index between the 2 groups (P<0.05) but not in age, gender, cement volume or volume-cubage index (P>0.05). Logistic regression analysis indicated that BMD, cement leakage and cement distribution index all significantly affected the occurrence of adjacent vertebral fractures following percutaneous vertebroplasty. CONCLUSION: A low BMD, cement leakage and a low cement distribution index are all risks factor of adjacent vertebral fracture after percutaneous vertebroplasty.

[p14ARF enhances cisplatin-induced apoptosis in human osteosarcoma cells in p53-independent pathway].

OBJECTIVE: To study effect of tumor suppressor p14ARF on cisplatin-induced apoptosis in human osteosarcoma cells with its molecular mechanisms to provide evidences for increasing chemosensitivity of osteosarcoma. METHODS: pcDNA3.1-p14ARF plasmid was stable transfected into MG63 cells lack of p14ARF expression. Expression of p14ARF on mRNA and protein level was evaluated with RT-PCR and Western blot. MG63, MG63-vec and MG63-ARF cells were treated with cisplatin. Cell growth inhibition and IC50 were determined through MTT assay. Apoptosis was detected using fluorescence-activated cell sorting and Hoechst33258 staining. The expression of p53, Bax, p21, Mdm2, Fas, Caspase-3, caspase-9 and PARP was detected with Western blot. RNAi was used to silence p53. Cells were pre-treated with Caspase-9 specific inhibitor Z-LEHD-FMK to determine whether the effect was Caspase-9-dependent. RESULTS: There was no expression of p14ARF in MG63 and MG63-vec cells but obvious expression in MG63-ARF cells on mRNA and protein level. Cell viability was 84.2%±4.3%, 80.8%±4.3% and 58.9%±5.4% in MG63, MG63-vec, and MG63-ARF cells after treatment of cisplatin for 72 h. IC50 was (11.1±0.6), (10.7±0.9) and (7.2±0.7) µmol/L. The apoptotic rate was 13.6%, 18.5% and 35.9% in groups, There were more obvious apoptotic more changes in MG63-ARF cells than MG63 and MG63-vec cells, and activation of Caspase-3, 9 and PARP on higher level in U2OS-ARF cells after stimulation with cisplatin for 72 h. The expression of p53, Bax, p21, Mdm2 and Fas, in MG63-vec and MG63-ARF cells did not changed (P>0.05). The expression of p53 was effectively and continuously suppressed by p53-siRNA in U2OS-vec and U2OS-ARF cells. The p53 silencing did not alter the cytotoxicity mediated by cisplatin treatment for 72 h (P>0.05). Cell viability was 96.8%±3.6%, 54.1%±5.7% and 89.5%±5.1% in Z-LEHD-FMK, cisplatin and Z-LEHD-FMK+cisplatin groups. CONCLUSION: p14ARF enhances cisplatin-induced apoptosis in human osteosarcoma MG63 cells in p53-independent caspase-9-dependent pathway, in which the intrinsic mitochondrial apoptotic pathway is involved.

[p14ARF enhances chemosensitivity to cisplatin in human osteosarcoma U2OS cells through p53 apoptotic pathway].

OBJECTIVE: To explore the effects of tumor suppressor p14ARF on chemosensitivity of human osteosarcoma U2OS cells to cisplatin and elucidate its molecular mechanism. METHODS: U2OS cells expressing no p14ARF and U2OS-ARF cells expressing p14ARF stably through stable transfection were treated with cisplatin. Cell viability and IC50 were assayed with methyl thiazolyl tetrazolium (MTT). Apoptosis was examined by fluorescence-activated cell sorting and Hoechst33258 staining. The expressions of p53, Bax, p21, Mdm2 and Fas were detected by Western blot. And colorimetry was used to determine the activities of caspase-3, caspase-8 and caspase-9. RESULTS: The viability was 84.8% ± 4.4%, 86.9% ± 5.0% and 66.7% ± 4.6% respectively in U2OS, U2OS-vec and U2OS-ARF cells. The values of IC50 were (15.8 ± 0.9) µmol/L, (16.3 ± 0.6) and (8.9 ± 0.8) µmol/L respectively in U2OS, U2OS-vec and U2OS-ARF cells. The levels of viability and IC50 obviously decreased in U2OS-ARF cells in response to cisplatin (P < 0.05). There were higher apoptotic rate and more obvious apoptotic morphological changes in U2OS-ARF cells than U2OS and U2OS-vec cells. The basal levels of p53, Mdm2 and p21 in U2OS-ARF cells were slightly higher than those in U2OS-vec cells. Cisplatin up-regulated p53, Mdm2 and p21 in both cell lines. However, the up-regulation was more pronounced in U2OS-ARF cells. Cisplatin did not change the levels of Bax and Fas in U2OS-vec cells. Bax protein was up-regulated in U2OS-ARF cells while the level of Fas remained constant. p14ARF also enhanced the activities of caspase-9 and caspase-3 in response to cisplatin. CONCLUSION: p14ARF enhances the chemosensitivity to cisplatin in human osteosarcoma U2OS cells through p53 apoptotic pathway. And intrinsic mitochondrial apoptosis is involved.

Biomechanical assessment of unilateral pedicle screws plus contralateral transfacetopedicular screws after transforaminal lumbar interbody fusion with two cages.

OBJECTIVE: To assess the biomechanical stability of unilateral pedicle screws (UPS) plus contralateral transfacetopedicular screws (TFPS) after transforaminal lumbar interbody fusion (TLIF) with two cages. METHODS: Range of motion (ROM) testing was performed in 28 fresh-frozen human cadaveric lumbar spine motion segments. The sequential test configurations included supplemental constructs after TLIF such as UPS, UPS plus contralateral TFPS and bilateral pedicle screws (BPS). All test specimens were fixated in the normal lordotic lignment, then mounted in a three-dimensional (3-D) motion testing machine and fixed to the load frame of a six degrees of freedom spine simulator. Each of the test constructs were subjected to three load-unload cycles in each of the physiologic planes generating flexion-extension, right-left lateral bending and right-left axial rotation load-displacement curves. Statistical analysis was performed on the ROM data. Comparison of data was performed by repeated-measures analysis of variance for independent samples followed by Bonferroni analysis for multiple comparison procedures. RESULTS: The ROMs for UPS, BPS and UPS plus TFPS fixation after TLIF were significantly smaller than those of the intact spine in all modes. The ROM for UPS plus TFPS fixation was between the largest for UPS and the smallest for BPS. The differences between ROMs of UPS and UPS plus TFPS were significant for both lateral bending and rotation. There were no significant differences between BPS and UPS plus TFPS in any mode. CONCLUSION: Because the UPS construct provides the least stability, especially during lateral bending and rotation, it should be used prudently. After TLIF with two cages, UPS plus TFPS provides stability comparable to that of TLIF with BPS. It is thus an acceptable option in minimally invasive surgery.

Effect of prenatal tetrandrine administration on transforming growth factor-beta1 level in the lung of nitrofen-induced congenital diaphragmatic hernia rat model.

PURPOSE: Tetrandrine (Tet) is a bisbenzylisoquinoline alkaloid isolated from the root of Stephania tetrandra, which has been used in traditional Chinese medicine to treat patients with silicosis, asthma, and pulmonary hypertension, and others and can be used as a pulmonary therapeutic agent. We hypothesized that it can also improve the lung growth in congenital diaphragmatic hernia (CDH) for its multiple biological effects. There are increasing evidences that suggest transforming growth factor beta1(TGF-beta1) plays a crucial role in fetal lung growth and morphogenesis. The aim of this study was to evaluate the effect of prenatal administration of Tet and to investigate its possible mechanism on the expression of TGF-beta1 in the lung of nitrofen-induced CDH rat model. METHODS: A CDH model was induced in pregnant Sprague-Dawley rats by administration of nitrofen on day 9.5 of gestation (Ed9.5 term, day 22). Tetrandrine (30 mg/kg) was given through gavage (once a day, for 3 days) on Ed11.5. Accordingly, there were 3 groups as follows: control (n = 9), CDH (n = 9), and CDH + Tet (n = 9). All the fetuses were delivered by cesarean delivery on Ed16.5, 18.5, and 21.5, respectively, to check if diaphragmatic hernia existed on each fetus, then the lung tissue weight (LW) and body weight (BW) of each fetus were recorded. Histologic evaluations and TGF-beta1 immunohistochemistry staining in the lung sample were performed for image analysis. RESULTS: Diaphragmatic hernia was observed in 95 of the 112 rat fetuses in CDH and CDH + Tet groups on Ed18.5 and Ed21.5 (84.8%), the incidence between the 2 groups had no statistical significance (P = .642). Lung weight/body weight in the CDH group and the CDH + Tet group were lower than that in the control group (P < .01), and LW/BW in the CDH group was lower than that in the CDH + Tet group (P < .05). Observed under the light microscope and electron microscope, marked hypoplasia of the lungs in fetuses among the CDH groups was observed, in contrast to improvement of the lungs in CDH + Tet fetuses. Statistical differences in morphological parameters (percentage of alveoli area, counting bronchus) were found even on Ed16.5 when diaphragm had not closed (P < .01). The number of type II pneumocytes and lamellar bodies in each group had no significant difference (P > .05). The immunoreactivity of TGF-beta1 in CDH group and CDH + Tet group were markedly stronger than that in the control group (P < .01). In addition, TGF-beta1 expression in the CDH group was stronger than that in the CDH + Tet group (P < .01). CONCLUSION: Nitrofen can interfere with lung development early in the fetal rat development before and separate from diaphragm development, and increased expression of TGF-beta1 in the lung of CDH rat model may suppress lung growth and development. Prenatal treatment with Tet can improve the growth of the lung of the nitrofen-induced CDH fetuses and its mechanism seems to be involved in downregulating the expression of TGF-beta1. It is a likely new approach to treat CDH and its coexistent lung hypoplasia by maternal Tet administration.

[A series of observation on the expression of TGF-beta1 in the lung of nitrofen-induced congenital diaphragmatic hernia rat model].

OBJECTIVE: To access the expression of transforming growth factor beta1 (TGF-beta1) in the lung of Nitrofen-induced congenital diaphragmatic hernia rat model. METHODS: Twelve timed-pregnant Sprague-Dawley rats were randomly divided into two groups, namely control group and CDH group on day 9.5 of gestation. Each rat in the CDH group was given 125 mg of Nitrofen (dissolved in seed fat) by gavage. Each rat in the control group was given the same dose of single oil. On day 16 of gestation, the two groups mentioned above were divided into three subgroups, and fetuses were delivered by cesarean section respectively on day 16, 18 and 21 of gestation. After the fetuses were checked for diaphragmatic hernia, lung tissue weight (LW) and body weight (BW) of each fetus on gestational day 21 were recorded. Lung histologic evaluations were made with microscope and TGF-beta1 immunohistochemistry staining were performed with image analyzing. RESULTS: At day 16 of gestation, no visible diaphragm closure was observed in all fetuses. Diaphragmatic hernia was observed in 32 of the 44 rat fetuses of the CDH groups on day 18 and day 21 of gestation (72.7%). Lw/Bw of the 21d subgroups of CDH group were lower than those of corresponding control group (P < 0.01). Observed under the microscope, the lungs of fetuses in CDH groups showed marked hypoplasia. The expression of TGF-beta1 was detected in cytoplasmic, without definite expression in nuclear. It was significantly stronger that the expression of TGF-beta1 was in the lungs of the CDH group than that of the control group (P < 0.01). CONCLUSIONS: Nitrofen interferes with lung development in early stage of the fetal before the diaphragm developed. TGF-beta1 would be one of the important factors which lead to pulmonary hypoplasia.

Prenatal tetrandrine treatment can reverse the abnormal conditions in the lung of newborn with congenital diaphragmatic hernia.

Pulmonary hypoplasia and persistent pulmonary hypertension are the most important reasons for the high morbidity and mortality of congenital diaphragmatic hernia (CDH). Despite surgical advances and advances in neonatal intensive care, the mortality still remains high. Then the research on how to improve prenatal fetal lung growth has become a focus. Some researches involved in fetal surgery, tracheal occlusion, prenatal use of corticosteroids etc., have been carried out in CDH animal models and humans. But the results either showed no benefit for the outcome of CDH or were unproved. Tetrandrine is a bisbenzylisoquinoline alkaloid isolated from the root of Stephania tetrandra. It has been used in traditional Chinese medicine for several decades to treat patients with silicosis, asthma and pulmonary hypertension etc. Some researches showed that prenatal tetrandrine administration can improve the lung development in CDH rat models. We hypothesize that prenatal treatment with tetrandrine can reverse the abnormal condition in the lung of newborn with CDH, and thus decrease the mortality.

Depressed exocytosis and endocytosis of type II alveolar epithelial cells are responsible for the surfactant deficiency in the lung of newborn with congenital diaphragmatic hernia.

Exocytosis and endocytosis are the way of macromolecules transmembrane transport. Pulmonary surfactant (PS), one of such macromolecules, is secreted via exocytosis of lamellar bodies and recycled via endocytosis by type II alveolar epithelial cells (AEC II). It maintains low alveolar surface tension and is therefore essential to normal lung function. PS deficiency causes respiratory distress syndrome in infants. Congenital diaphragmatic hernia is an abnormal condition in which low lung compliance is involved. This condition is multifactorial and a primary surfactant deficiency may be responsible for it. We hypothesize that surfactant deficiency is involved in CDH and depressed activity of exocytosis and endocytosis in AEC II is responsible for the surfactant deficiency in the lung of newborn with congenital diaphragmatic hernia.

[A morphological study with serial histological slices on the normal and abnormal gubernacula in newborn male mice].

OBJECTIVE: To explore the feasibility of serial slices microscopic histological investigation for the elaborate evaluation of reproductive system malformations. METHODS: Newborn male mice prenatally exposed to different doses of subcutaneously given diethylstilbestrol (DES) from gestational day 9 to 17 were treated by fixing parts of the abdomen in situ and setting them to transected serial slices. All the slices were stained, studied under the microscope and serially recorded by software. The gubernaculum was morphologically analyzed and its location and size were measured. RESULTS: Morphologically, the gubernaculum could be identified clearly, its structure inhomogeneous from proximal to distal and dissymmetric from right to left. The environmental estrogen produced different effects on the morphology of the gubernaculum in different parts and most obviously affected its length. CONCLUSION: Prenatal exposure to environmental estrogen has evident and general effects on the gubernacular development of newborn male mice. The morphological study with serial histological slices gives a precise and systematic evaluation of genital malformations.

Interferonalpha enhances etoposide-induced apoptosis in human osteosarcoma U2OS cells by a p53-dependent pathway.

Interferonalpha (IFNalpha) induces cell cycle arrest and triggers apoptosis and chemosensitivity. But the mechanism of IFNalpha in regulating chemosensitivity has not been fully understood. To study whether IFNalpha affected chemosensitivity of osteosarcoma cells, we treated p53-wild U2OS cells and p53-mutant MG63 cells with IFNalpha and etoposide, alone or in combination, and then examined growth inhibition, cell cycle arrest and apoptosis. IFNalpha enhanced etoposide-induced growth inhibition and apoptosis in p53-wild U2OS cells but not p53-mutant MG63 cells in a dose- and time-dependent manner. Etoposide-induced G2/M phase arrest was also enhanced by IFNalpha. The enhanced apoptosis was associated with the accumulation of transcriptionally active p53 accompanied with increased Bax and Mdm2, as well as decreased Bcl-2. IFNalpha also activated caspases-3, -8 and -9 protein kinases and PARP cleavage in response to etoposide in U2OS cells. Moreover, the combination-induced cytotoxicity and PARP cleavage were significantly reduced by caspase pan inhibitor and p53 siRNA. Thus we conclude that IFNalpha enhances etoposide-induced apoptosis in human osteosarcoma U2OS cells by a p53-dependent and caspase-activation pathway. The proper combination of IFNalpha and conventional chemotherapeutic agents may be a rational strategy for the treatment of human osteosarcoma with functional p53.

[Effect of tetrandrine on endothelin expression in the lungs and clinical significance thereof: experiment with rat models with nitrofen-induced congenital diaphragmatic hernia].

OBJECTIVE: To investigate the effects of tetrandrine on endothelin expression in the lungs and its clinical significance. METHODS: 25 pregnant female SD rats were randomly divided into 5 equal groups: Groups A to D were fed with nitrofen to cause congenital diaphragmatic hernia (CDH) in the fetuses. Group A was injected with normal saline, Group B with dexamethasone (DXM), Group C with tetrandrine, and Group D with DXM + tetrandrine. Group E was control Group. On day 21.5 of pregnancy the fetuses were delivered by cesarean section and killed. Microscopy was used to observe the CDH formation, and the relative wall thickness (RWA) and relative wall area (RWA) of pulmonary arterioles. The lung/body weight ratio, and relative integrated optical density (IOD) of pulmonary arteriole and bronchiole were observed. The expression of endothelin in the lung tissues was detected by immunohistochemistry. RESULTS: 9 rats of Group A-D produced 57 fetuses with CDH with a CDH arte of 45.2%. The lung/body weight ratios, and RWA values of Group A-D were all significantly lower than that of Group E (all P < 0.05). The RWT of pulmonary arteriole was significantly lower in Groups B and C compared with Group E (both P < 0.05). The RWT and RWA of Group A were significantly lower than those of Group B-D (all P < 0.05). The values of relative IOD of pulmonary tissues and of pulmonary arteriole of Group A-D were all significantly lower than those of Group E (all P < 0.05). A positive correlation existed between the relative IOD of endothelin in pulmonary arteriole and in bronchiole (P < 0.01), and among the RWT and RWA of pulmonary arteriole, and relative IOD (all P < 0.01). CONCLUSION: Tetrandrine improves the pulmonary hypoplasia and degrades the pulmonary hypertension.

Expression of glucocorticoid receptor isoforms in cutaneous hemangiomas and vascular malformations.

BACKGROUND: Hemangiomas are the most common tumors in children. Some hemangiomas may require intervention because of their location, size, behavior, or potential for important complications. Pharmacological therapy with glucocorticoids is the mainstay treatment, but there is no consensus on therapeutic regimens or candidate selection, therapeutic efficacy varies, and the mechanism mediating the beneficial effects of glucocorticoids remains unclear. This study was performed to investigate the expression patterns of the glucocorticoid receptor (GR) and its alpha isoform (GRalpha) in cutaneous hemangiomas and vascular malformations. METHODS: SP immunohistochemical technique was used to examine the expression of GR(e-20) (GR) and GR(p-20) (GRalpha) on vascular endothelial cells in 80 specimens that included 33 proliferating hemangiomas, 32 involuting hemangiomas, 7 vascular malformations as well as 8 normal skin tissues, all obtained from infants and children. GR and GRalpha expression in prepared tissue slides were examined using automated computer-assisted microscopic analysis. Mean gray scale values were compared among the various tumor types. RESULTS: The mean gray scale values of GR were 127.0 +/- 6.4 and 121.4 +/- 6.6 in hemangiomas and vascular malformations respectively, but this difference was not statistically significant (P = 0.104). However, these values were all markedly higher than that of normal skin, which was only 108.6 +/- 6.8 (P = 0.001 and P = 0.000 for comparison with hemangiomas and vascular malformations respectively). The gray scale of GR in proliferation and involuting hemangiomas were 127.9 +/- 4.8 and 126.0 +/- 5.8 respectively, but this difference was not significant (P = 0.146). However, GRalpha expression in hemangiomas, vascular malformations and normal skin declined gradually in stepwise fashion (127.3 +/- 5.4, 120.4 +/- 6.1 and 109.9 +/- 5.3 respectively; P < 0.001). GRalpha expression was higher in proliferating hemangiomas than in involuting hemangiomas (127.2 +/- 6.3 and 122.5 +/- 6.3; P = 0.004). CONCLUSIONS: GR and GRalpha are strongly expressed in hemangiomas and vascular malformations. The expression of GRalpha is closely related to the phase of the hemangioma. Determination of GR and GRalpha may be a positive significance to understand the information of hemangiomas and vascular malformations and may further help determining proper strategies of steroid therapy for hemangiomas and vascular malformations.