Kinesiska forskare närmare Gliomaeländets lösning

12 kinesiska forskare har genomfört en studie om den riktigt eländiga formen av hjärncancer,glioma
Denna cancertyp som har så dålig prognos för de drabbade.Därför ska dessa kinesiska forskare honoreras för sina ansträngningar att hitta botemedel mot denna best.Naturligtvis har forskarna haft HoloMonitor vid sin sida för att åstadkomma de resultat denna studie visar upp.

Wiki berättar mera om denna cancertyp :
- Gliom är en tumör som liknar gliaceller i centrala nervsystemet. Det är den vanligaste hjärntumören hos vuxna.Benämningar som oligodendrogliom, oligoastrocytom, eller astrocytom anger vilken celltyp gliomet liknar. Dock finns också sentida teorier att gliomen uppkommer genom förändringar i stamceller och liknande celler, i nervsystemet. Den senare teorin är kontroversiell men vinner allt större acceptans. Tidigare förelåg konsensus om att astrocyter är cellursprung till gliomen.
Glioblastom är en mycket aggressiv tumörtyp som bland annat leder till nekros. Den indelas i primär typ och sekundär typ. Den primära typen, som vanligen drabbar äldre, bryter ut snabbt och utan föregående konstaterade lågmaligna tumörer. Sekundär typ bryter oftast ut före 45 års ålder från mera godartade tumörer och har ett långsammare förlopp. Båda typerna har vanligen mutationer i generna för tumörprotein 53, PTEN, och epidermala tillväxtfaktorns receptor. De maligna gliomen påverkar immunsystemet.

De kinesiska forskarna har som utgångspunkt haft just att utgå från nervsystemet.
Men till deras forskningsrapport som publiceras av det vetenskapliga organet Clinical and Translational Medicine.

SUMOylation of PUM2 promotes the vasculogenic mimicry of glioma cells via regulating CEBPD

06 September 2020

Abstract

Glioma is the most common form of primary central nervous malignant tumors. Vasculogenic mimicry (VM) is a blood supply channel that is different from endothelial blood vessels in glioma. VM is related to tumor invasion and metastasis. Therefore, it plays an important role to target therapy for glioma VM. Our experimental results showed abnormal expression of UBE2I, PUM2, CEBPD, and DSG2 in glioma cells. The Co‐IP and Immunofluorescence staining were used to detect that PUM2 can be modified by SUMO2/3. The interaction between PUM2 and CEBPD mRNA was detected by the RIP assays. The interaction between transcription factor CEBPD and promoter region of DSG2 was detected by the ChIP assays and luciferase assays. The capacity for migration in glioma cells was observed by the laser holographic microscope. The capacity for invasion in glioma cells was detected by Transwell method. The VM in glioma cells was detected by three‐dimensional cell culture method. The experimental results found that the upregulation of UBE2I in glioma tissues and cells promotes the SUMOylation of PUM2, which decreases not only the stability of PUM2 protein but also decreases the inhibitory effect of PUM2 on CEBPD mRNA. The upregulation of CEBPD promotes the binding to the upstream promoter region of DSG2 gene, further upregulates the expression of DSG2, and finally promotes the development of glioma VM. In conclusion, this study found that the UBE2I/PUM2/CEBPD/DSG2 played crucial roles in regulating glioma VM. It also provides potential targets and alternative strategies for combined treatment of glioma.

Ur studien tar jag mig friheten att klistra in de delar där man använt sig av HoloMonitor.(Fetningar och understrykningar är som vanligt mina egna.)

2.8 Cell migration assays

The capacity for migration in glioma cells was observed by the HoloMonitor M4 culture system (Phase Holographic Imaging PHI AB, SE) in vitro. For details of the experiment, please refer to online Additional Materials and Methods.
Cell migration assays was performed using the HoloMonitor M4 culture system
(Phase Holographic Imaging PHI AB, SE) according to the manufacturer’s protocols.
The cells of each group were inoculated into a six-well plate at a concentration of 2×104cells/ml. After the cells were attached to the petri dish, they were placed on the HoloMonitor M4 culture system and set for imaging for 6h at 1h  intervals.For each experimental group,  we show the last image frame  and the cell movements.  At the start of the analysis 5 visually identifiable cells in each experimental set were selected for tracking. Their movements are displayed in spatial X-Y plots.

3.2 UBE2I promoted the capacities for migration, invasion, and VM in glioma cells

Finally, the invasion, migration, and VM capacities were also analyzed by Hstudio M4 software, transwell assay, and 3‐D cell culture, respectively. As shown in Figure 2F‐H, the knockdown of UBE2I significantly inhibited the capacities for migration, invasion, and VM in glioma cells, while the overexpression of UBE2I had the opposite effects (online Additional Figure 2). These results suggest that UBE2I regulates the expression of PUM2 in glioma cells, thereby further regulating the capacities for migration, invasion, and VM in glioma cells. 

(F) The Hstudio M4 system observed the capacity for migration in the UBE2I knockdown cells (U251 and U373), (n = 5). Scale bars: 100 µm. (G) Transwell method was used to detect the capacity for invasion in the UBE2I knockdown cells (U251 and U373). Scale bars: 100 µm. (H) Three‐dimensional cell culture method was used to detect the change of VM in the UBE2I knockdown cells (U251 and U373). Scale bars: 200 µm. Each value represents the mean±SD (n = 3), *P < .05, **P < .01, compared with sh‐NC group.

3.3 UBE2I mediated the capacities for migration, invasion, and VM in glioma cells by regulating the expression of PUM2

Using the Hstudio M4 system, Transwell method, and three‐dimensional cell culture method, it was found that PUM2 overexpression significantly inhibited the capacities for migration, invasion, and VM in glioma cells compared with the NC group. Our experimental results also reported that UBE2I overexpression rescued the effect of PUM2 overexpression on the capacities for migration, invasion, and VM in glioma cells (Figure 3E‐G). Therefore, it was confirmed that UBE2I decreased the expression of PUM2 protein by SUMOylation in glioma cells, which further inhibited the capacities for migration, invasion, and VM in glioma cells.

(E) Hstudio M4 system was used to observe the change of the capacity for migration in the cells (U251 and U373) overexpressed UBE2I after overexpressed PUM2 (n = 5). Scale bars: 100 µm.

3.4 PUM2 inhibited CEBPD expression by binding CEBPD mRNA, thereby inhibiting the capacities for migration, invasion, and VM in glioma cells

Using the Hstudio M4 system, the Transwell method, and the three‐dimensional cell culture method, it was found that CEBPD knockdown significantly inhibited the capacities for migration, invasion, and VM in glioma cells compared with the sh‐NC group (Figure 4G‐I). The results confirmed that the UBE2I/PUM2 axis regulated the expression of CEBPD. Compared with HA cells, CEBPD expression in glioma cells was significantly increased, and CEBPD knockdown significantly inhibited the capacities for migration, invasion, and VM in glioma cells.

(G) Hstudio M4 system was used to observe the change of the capacity for migration in cells (U251 and U373) CEBPD knockdown (n = 5). Scale bars: 100 µm. (H) Transwell method was used to detect the change of the capacity for invasion incells (U251 and U373) CEBPD knockdown. Scale bars: 100 µm. (I) Three‐dimensional cell culture method was used to detect the change of VM in the cells (U251 and U373) CEBPD knockdown. Each value represents the mean±SD (n = 3), **P < .01, ***P < .001, compared with sh‐NC group

3.5 CEBPD promoted the capacities for migration, invasion, and VM in glioma cells by regulating the transcription of DSG2

Through the Hstudio M4 system, Transwell method, three‐dimensional cell culture method, it was found that DSG2 knockdown significantly inhibited the capacities for migration, invasion, and VM in glioma cells compared with the sh‐NC group, and CEBPD overexpression significantly rescued the effect of DSG2 knockdown (online Additional Figure 3A,B and Figure 5G).
This study confirmed that CEBPD promoted the expression of DSG2 by transcription in glioma cells, thereby promoting the capacities for migration, invasion, and VM in glioma cells.

Längst ner på sidan under fliken Supporting Information hittar ni mer info om användande av det excellenta instrumentet HoloMonitor.

Läs även gärna hela slutstycket Discussions.Jag klipper in del av det.Betänk att utan PHI,s teknik hade man inte kommit fram till dessa resultat.

The expression of UBE2I in glioma cells is significantly increased, thereby promoting PUM2 SUMOylation, leading to the degradation of PUM2 protein by proteasome. UBE2I inhibits the role of PUM2 protein in the degradation of CEBPD mRNA. CEBPD overexpression promotes the transcriptional expression of DSG2, which in turn promotes the capacities for migration, invasion, and VM in glioma cells.

In summary, this study first discovered the abnormal expression of UBE2I, CEBPD, PUM2, and DSG2 in glioma tissues and cells. UBE2I knockdown, CEBPD knockdown, and PUM2 overexpression significantly inhibited the capacities for migration, invasion, and VM in glioma cells. UBE2I knockdown inhibited PUM2 SUMOylation, resulting in increased stability of PUM2 protein and increased expression of PUM2 protein in the cytoplasm. Further, PUM2 overexpression bound to the PRE of CEBPD mRNA and promoted the degradation of CEBPD. CEBPD knockdown inhibited the transcriptional regulation of the DSG2, thereby inhibiting the capacities for migration, invasion, and VM in glioma cells. The results of this study confirmed that UBE2I, CEBPD, and PUM2 have the potential to be new molecular targets, and the research on the treatment of gliomas needs to be further explored.

Min kommentar

Än en gång visar forskare upp helt nya resultat med stöd av HoloMonitor, insikter och kunskap som kommer gagna alla cancerdrabbade. Undertecknad har sagt det förut,men jag upprepar mig igen:

Utrusta världens alla cancerforskare med varsitt HoloMonitor instrument! Ge besten en match och jämna ut oddsen till fromma för cancerdrabbade.

De kinesiska forskarna ska ha stor cred för deras engagemang att hitta lösning för alla gliomadrabbade.

                                        Mvh the99