Introduction
The traditional Chinese medicine strategy for body wellness and
longevity focused on the balancing of Yin and Yang, or Xue (blood,
nutrient) and Qi (energy). This anti-aging strategy applies to the
prescription of Chinese medicine and generates varies classical
prescriptions for treatment of aging-related disease and sustaining
longevity. One of the most extensively used prescriptions for longevity
is the Bazhen decoction. Bazhen decoction is composed of eight herbal
medicine, including
Panax
ginseng, Radix rehmanniae praeparata, Radix paeoniae
alba, Ligusticum wallichii,
Poria cocos, Angelica
sinensis, Rhizoma Atractylodis Macrocephalae, and
Licorice. Bazhen decoction is the combined prescription of Sijunzi and
Siwu decoction, and is famous for enhancing both Qi and Xue, and
balancing Yin and Yang. Thus, it is the one of the extensively used
prescription for the intervention of sub-health status, and the
prevention of aging-related degenerative diseases [1-4].
It has been revealed that Bazhen decoction was effective in the
treatment of 5-fluorouracil-induced anemia in mice. It could promote the
proliferation and differentiation of bone marrow cells, elevate the
transcription of EPO mRNA, increase the red blood cell count and the
hemoglobin concentration [5]. The similar effect of Bazhen has been
found in the bone marrow depression induced by cyclophosphamide in mice,
by promoting the proliferation of hematopoietic progenitor cell and
secretion of hematopoietic growth factor [6]. It has also been shown
that Bazhen decoction combined with sequential treatment of chemotherapy
on acute lymphoblastic leukemia patients with deficiency of Qi and Yin
could protect the hematopoietic system from damages induced by
chemotherapy, and decrease the incidence of nausea and vomiting, liver
and kidney injury [7]. Together these data suggest the function of
Bazhen decoction in promoting hematopoietic regeneration.
It has also been found that Bazhen decoction administration could
decrease acetaminophen-induced liver injury marker, maintain the
activity of anti-oxidative factors, and depress the expression of
pro-inflammatory factors. These data suggest that Bazhen decoction
protect against acetaminophen induced acute liver injury by inhibiting
oxidative stress, inflammation and apoptosis in mice [8]. The
modification of Bazhen decoction (Huangqi Bazhen decoction) has been
used to interfere the chemotherapeutic intestinal mucositis induced by
capecitabine in mice. The results showed that the intestinal injury
(villus height shortening, crypt destruction, and apoptosis) was
reversed by Bazhen decoction, and the weight loss and diarrhea during
capecitabine treatment were reduced [9]. These data suggest that
Bazhen decoction could prevent tissue damages induced by toxic
chemicals, such as chemotherapy drugs. The Bazhen decoction has been
also apply to the treatment of premature ovarian failure induced by
D-galactose in rats, it could up-regulate the E2 and X-linked inhibitors
of apoptosis protein (XIAP), and prevented the apoptosis of oocyte and
granulosa cells [10].
Together these data reveal the promising application of Bazhen decoction
in prevent tissue damage and aging-related tissue degeneration. However,
due to the complicity of the decoction components, the full image of the
pharmacological basis of Bazhen decoction is still unclear, which hinder
the precise clinical application, especially for complicated aging
related degenerative diseases. The further dissection of the molecular
pathways regulated by Bazhen decoction might provide the molecular
pathways for the evaluation of treatment effect, and provide readouts
for more precise symptoms and timing for the clinical application of
Bazhen decoction on aging related diseases.
The hallmarks of aging include the attenuation of stem cell capacity,
abnormal intercellular communication, abnormal mitochondrial function,
increased chromosomal instability, altered epigenetic modifications,
decreased protein homeostasis, decreased telomerase activity, and
abnormal telomere function [11].
Telomere is at the end of eukaryotic chromosomes and composed of repeat
DNA sequence TTAGGG and shelterin protein complex. Telomere is essential
in maintaining the integrity of chromosomal DNA, and preventing
chromosome ends from being recognized as DNA damage [12, 13].
Telomere length is mainly maintained by telomerase, or by a class of
mechanisms referred to as alternative lengthening of telomere (ALT)
[14]. In the absence of telomere lengthening mechanism, telomere DNA
will gradually shorten along with the cell division, the shortened
telomere activates DNA damage responses, such as p53-regulated signaling
and the inflammation response pathways, eventually induced cellular
senescence, apoptosis, and organism progeroid diseases [15].
In addition to telomerase, DNA helicases are also required for proper
replication and elongation of telomeric DNA. Telomeric DNA is rich in G
and tends to forms G-quadruplex DNA (G4 structure), which requires
helicases to unwind properly for further DNA replication [16]. DNA
helicases are a large family of proteins that hydrolyze ATP to produce
energy to unwind DNA double helix. The RecQ family of DNA helicases,
such as Wrn, Blm, Recql4, are involved in DNA replication,
recombination, DNA damage repair and telomere maintenance, and play an
essential role in maintaining chromosome stability [17]. Thus, the
function of DNA helicase improves the G4 DNA unwinding, facilitates the
DNA replication and the telomere elongation.
It has been proved by omics and large sample population studies that
telomere length gradually decreases with human aging, and interacts with
other important aging markers (such as stem cell function, mitochondrial
function, and immune function), leading to a gradual decline in tissue
renewal ability and inflammation. Telomere length has become one of the
gold standards for detecting the process of aging and its related
diseases [18-20].
To explore the full image of the molecular pathways that Bazhen
decoction regulates, in this study, we adopted the RNA sequencing
technique to dissect the molecular pathways regulated by Bazhen
decoction. RNA sequencing could reveal all the RNAs that expressed under
certain circumstance, thus provide the full image of transcriptome, in
this case, the cellular transcriptome induced by Bazhen decoction
treatment. By further analysis of the transcriptome, we found the DNA
replication and telomere maintenance function been improved by Bazhen
decoction treatment. To verify these data, we utilize the mouse embryo
fibroblasts (MEFs) generated from Werner syndrome mouse model. Werner
syndrome (WS) is an autosomal recessive genetic disease caused by DNA
helicase Wrn gene mutation and telomere dysfunction. WS is characterized
by premature aging and shortened life span. The average lifespan of WS
is only 46-48 years, the symptoms include premature atherosclerosis,
osteoporosis, cataracts, reproductive deficiency, type 2 diabetes
mellitus, and soft-tissue sarcoma. We revealed that the Bazhen decoction
elevated the expression of DNA helicases, which promoted the G4 DNA
resolving, facilitated the DNA replication and the telomere elongation,
and facilitate the progeroid Werner syndrome cell proliferation.