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The mammalian genome transcribes several thousand incRNAs but how incRNAs function is just being

understood. So, mechanisms of action for only a few are described. Evidence suggests that incRNAs may
contribute to control of imunity, tirmorigenesis, cell proliferation, differentiation and apoptosis.
However, systematic identification and characterization of incRNAs involved in hepatitis viral infection
and hepatic lipid metabolism are lacking.

We repot the discovery of a novel incRNAs molecule incHR1, which suppresses the SREBP-1 c expression
and partipates in lipid metabolism regulation. Our data suggest that TG and LD accumulation can be
inhibited by incHR1 in both cultured hepatoma cells and in transgenic mice.

Genom mamalia mentranskripsi beberapa ribu incRNAs tapi bagaimana fungsi incRNAs hanya dipahami.
Jadi, mekanisme aksi untuk hanya beberapa dijelaskan. Bukti menunjukkan bahwa incRNAs dapat
berkontribusi untuk kontrol imunity, tirmorigenesis, proliferasi sel, diferensiasi dan apoptosis. Namun,
identifikasi sistematis dan karakterisasi incRNAs terlibat dalam infeksi virus hepatitis dan metabolisme
lipid hati kurang.

Kami merepoting penemuan sebuah novel incRNAs molekul incHR1, yang menekan SREBP-1 c
berekspresi dan partipates dalam peraturan metabolisme lipid. Data kami menunjukkan bahwa TG dan
akumulasi LD dapat dihambat oleh incHR1 di kedua sel hepatoma berbudaya dan pada tikus transgenik.

Figure 2

Biological characteristics if incHR1 . (A) structural diagram of incHR1 gene. (B) Endogenous incHR1
expression was measured in 11 cell lines using absolute qRT-PCR. (C) intracellular localization of incHR1
was visualized in Huh7 cells with RNA-FISH assays (blue for nucleus and green for incHR1) .scale bars ,
13um.

Gambar 2

karakteristik biologi jika incHR1. (A) diagram struktural gen incHR1. ekspresi (B) endogen incHR1 diukur
dalam 11 baris sel menggunakan absolut QRT-PCR. (C) lokalisasi intraseluler dari incHR1 divisualisasikan
dalam sel Huh7 dengan tes RNA-FISH (biru untuk inti dan hijau untuk incHR1) .scale bar, 13um.

Figure 3

LncHR1 negatively regulates endogenous SREBP-1c and FAS. (A) LncHRI 1 overxpression plasmid was
transfected into Huh7 cells in a dose dependent manner and intracellular LncHR was quantified by qRT-
PCR. An empty vector was a negative control. The data were normalized to relative fold changes. B) post
transfection of LncHRI overexpression plasmid for 48h, endogenous SREBP-1c mRNA (up) and protein
(down) were measured by qRT-PCR and western blot, respectively. C) LncHRI knockdown plasmid
(shRNA) was transfected into Huh7 cells in a dose dependent manner and intracellular LncHRI lever was
quantified by qRT-PCR. D) forty-eight hours post transfection LncHRI knockdown plasmid, endogenous
SREBP-1c mRNA(up) and protein (down) levels were measured by qrt-pcr and western blot, respectively.
E) hcv was incolated into huh7 cells 24 before transfection with either control vector or LncHRI
Overexpression plasmid and 48 h post-transfection, endogenous SREBP-1 c protein was measured with
western blot. Actin was used as an equal loading control. F) endogenous FAS protein expression was
measured in huh7 cells with overexpressed (up) or decreased (down) LncHRI

. empty vectors were negative controls. Data are means +- SD p<0.005, p<0,001

LncHR1 negatif mengatur endogen SREBP-1c dan FAS. (A) LncHRI 1 overxpression plasmid itu transfected
ke sel Huh7 dengan cara yang tergantung dosis dan LncHR intraseluler dikuantifikasi dengan QRT-PCR.
Vektor kosong adalah kontrol negatif. Data yang dinormalisasi dengan perubahan kali lipat relatif. B)
pasca transfeksi LncHRI berlebih plasmid untuk 48 jam, endogen SREBP-1c mRNA (atas) dan protein
(bawah) diukur dengan QRT-PCR dan blot Barat, masing-masing. C) LncHRI knockdown plasmid (shRNA)
telah transfected ke sel Huh7 dengan cara yang tergantung dosis dan intraseluler LncHRI tuas
dikuantifikasi dengan QRT-PCR. D) empat puluh delapan jam pasca transfeksi LncHRI knockdown
plasmid, endogen SREBP-1c mRNA (atas) dan protein (bawah) tingkat diukur dengan QRT-pcr dan blot
Barat, masing-masing. E) HCV incolated ke dalam sel huh7 24 sebelum transfeksi dengan baik vektor
kontrol atau LncHRI

Ekspresi plasmid dan 48 jam pasca-transfeksi, endogen SREBP-1 c protein diukur dengan blot barat.
Aktin digunakan sebagai kontrol pemuatan sama. F) endogen ekspresi protein FAS diukur dalam huh7
sel dengan diekspresikan (up) atau menurun (down) LncHRI

. vektor kosong yang kontrol negatif. Data adalah sarana + - SD p <0,005, p <0,001

Figure4

LncHRI supessed tg accumulation and ld formation in steatotic hepatocytes. Huh7 cells were transfected
with LncHRI overexpression A) or knockdown B) plasmids, and then treated with oa at 24h post
transfection.an empty vector was used as negative control. Endogenous SREBP-1c protein was
measured with western blot (A,B) and intracellular TG was quantified (C,D) . intracellular LD formation in
huh 7 cells was visualized with oil red o staining. The nuclei were indicated by DAPI staining. Scale bars,
30 um (E,F). volume of LD in each treatment was determined with image j software (G,H) . data are
means +- SD p<0,005 . P<0.001

Genomic studies have identified numerous genes involved in the lipid metabolism that ca be affected by
HCV infection. Currently, few studies about the function of liver disease related LncRNA, LncLSTR which
regulates clearance during lipid homeostasis. However, the relationships of LncRNAs associated with
hepatitis viral infection and hepatic lipid metabolism in human cells are unknown. Interestingly, our
efforts to screen HCV infection-regulated hosts LncRNAs revealed that LncHRI is anegative regulator of
SREBP -1c expression and tg accumulation. Alignment sequence analysis indicated that LncHRI is a
human specific LncRNA. To our knowledge, LncHRI is the first LncRNA reported to regulate lipid
synthesis via SREEP-1c.

Hepatic lipid abronarmalities are a characteristic feature of HVC infection. In experimental studies using
cultured cells, the expression of transfected HCV core proteins has been found to increase lipid
accumulation . studies examining the effect of hcv invection on srebp-1c have shown that the genes are
transcriptionally induced, and their proteolytic cleavage is stimulated. Then, in another study, the
expression of hcv core protein was found to activate the FAS. Our results indicated that LncHRI inhibited
the expression of srebp-1c by hcv infection. The suggest that LncHRI is able to inhibit viral infection and
protect the host

Figure4

LncHRI ditekan akumulasi tg dan pembentukan ld dalam hepatosit steatotic. Sel-sel Huh7 transfected
dengan LncHRI berlebih A) atau knockdown B) plasmid, dan kemudian diobati dengan oa di 24h pasca
transfection.an vektor kosong digunakan sebagai kontrol negatif. Endogen protein SREBP-1c diukur
dengan western blot (A, B) dan intraseluler TG dikuantifikasi (C, D). pembentukan LD intraseluler di huh
7 sel divisualisasikan dengan minyak merah o pewarnaan. Inti yang ditunjukkan oleh DAPI pewarnaan.
Skala bar, 30 um (E, F). Volume LD di setiap perlakuan ditentukan dengan software image j (G, H). Data
adalah sarana + - SD p <0.005. P <0,001

Studi genomik telah mengidentifikasi banyak gen yang terlibat dalam metabolisme lipid yang ca
terpengaruh oleh infeksi HCV. Saat ini, beberapa studi tentang fungsi penyakit hati terkait LncRNA,
LncLSTR yang mengatur izin selama homeostasis lipid. Namun, hubungan LncRNAs terkait dengan infeksi
virus hepatitis dan metabolisme lipid hati dalam sel manusia tidak diketahui. Menariknya, upaya kami
untuk menyaring HCV infeksi-diatur host LncRNAs mengungkapkan bahwa LncHRI adalah anegative
regulator dari SREBP 1 C akumulasi ekspresi dan tg. analisis urutan keselarasan menunjukkan bahwa
LncHRI adalah LncRNA tertentu manusia. Untuk pengetahuan kita, LncHRI adalah LncRNA pertama kali
dilaporkan untuk mengatur sintesis lipid melalui SREEP-1c.

abronarmalities lipid hati adalah fitur karakteristik infeksi HVC. Dalam penelitian eksperimental
menggunakan sel kultur, ekspresi protein inti HCV transfected telah ditemukan untuk meningkatkan
akumulasi lipid. penelitian yang meneliti efek HCV invection pada SREBP-1c telah menunjukkan bahwa
gen transcriptionally diinduksi, dan pembelahan proteolitik mereka dirangsang. Kemudian, dalam studi
lain, ekspresi protein HCV inti ditemukan untuk mengaktifkan FAS. Hasil kami menunjukkan bahwa
LncHRI menghambat ekspresi SREBP-1c oleh infeksi HCV. The menyarankan bahwa LncHRI mampu
menghambat infeksi virus dan melindungi host